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Glucosa-6-fosfato deshidrogenasa (G6PD). Respuesta de los hematies y otras celulas humanas a la disminucion en su actividad.

RESUMEN

La glucosa-6-fosfato deshidrogenasa (G6PD) es la primera enzima de la via pentosa fosfato y la principal fuente intracelular de nicotidamina adenina dinucleotido fosfato reducido (NADPH), compuesto comprometido en diversos procesos fisiologicos, por ejemplo defensa antioxidante (sobre todo celulas como los eritrocitos), modulacion del crecimiento endotelial, eritropoyesis, vascularizacion y fagocitosis. La deficiencia de G6PD es la enzimopatia ligada al cromosoma X mas comun en el ser humano. Si bien se puede presentar en cualquier tipo de celula, su carencia absoluta es incompatible con la vida. Segun la OMS, en el mundo hay mas de 400 millones de personas afectadas por la deficiencia de la enzima, y para Colombia calculan una prevalencia de la deficiencia severa entre 3% y 7%, pero no se conocen los datos relativos a las alteraciones leves y moderadas, que tambien tienen efectos clinicos. El presente articulo revisa los aspectos biomoleculares mas importantes de la enzima, su clasificacion de acuerdo con la actividad y la movilidad electroforetica, y tambien se mencionan algunos aspectos clinicos relacionados con la alteracion de su actividad.

Palabras clave: Ultraestructura; Fisiologia; Genetica; Epidemiologia; Deficiencia de glucosa-6-fosfato deshidrogenasa; Eritrocitos; Anemia hemolitica congenita.

Glucose-6-phosphate dehydrogenase (G6PD). Response of the human erythrocyte and another cells to the decrease in their activity

SUMMARY

Glucose-6-phosphate dehydrogenase is the first enzyme in the pentose phosphate pathway and the main intracellular source of reduced nicotidamineadenine nucleotidephosphate (NADPH), involved in diverse physiological processes such as antioxidant defense, (for instance in the erythrocyte) endothelial growth modulation, erithropoyesis, vascularization and phagocitosis. G6PDH deficiency is the most common X-chromosome-linked enzymopathy in human beings. Although it is present in any type cell, its absolute deficiency is incompatible with life. According to WHO, 400 million people are affected by G6PD deficiency in the world but in Colombia, the severe form prevalence is about 3% to 7%. There are no data related to slight and moderate alterations, that also have clinical effects. This paper reviews some G6PD biomolecular aspects, its classification according to activity and electrophoretic mobility, as well as some main clinical aspects related to its activity alteration.

Keywords: Erythrocyte; Physiology; Genetics; Epidemiology; Glucose-6-phosphate dehydrogenase deficiency; Congenital hemolytic anemia.

Todos los organismos vivientes, sean levaduras o protozoos, plantas o animales, expresan la enzima glucosa- 6-fosfato deshidrogenasa (G6PD) (1).

Aunque la G6PD se encuentra en el citoplasma de todas las celulas de los mamiferos, en los globulos rojos su deficiencia es mas evidente, quiza porque estas celulas viven durante largo tiempo sin nucleo y porque contienen proteasas que degradan la enzima mutante en mayor grado de lo que hacen las proteasas de otros tejidos1.

Como el eritrocito es una celula transportadora de oxigeno por excelencia sus mecanismos de defensa frente al estres oxidativo hacen parte del mantenimiento de este en circulacion. Estos mecanismos de defensa dependen en gran parte del suministro metabolico de la forma reducida de NADP (NADPH + [H.sup.+]). Debido a las caracteristicas metabolicas particulares de estas celulas, tan solo las dos primeras reacciones de la via de las pentosas (tambien llamada de la hexosa monofosfato) tienen la capacidad de generar NADPH + H. Estas son primero, la conversion de glucosa-6-fosfato en acido-6-fosfogluconico y segundo, la conversion de este intermediario en ribulosa- 5-fosfato con desprendimiento de C[O.sub.2]. Las dos reacciones son secuenciales y en ambas el NADP es reducido. Mientras que la primera es catalizada por la enzima G6PD, la segunda lo es por la 6-fosfogluconato deshidrogenasa. Mediante la produccion del NADPH los eritrocitos reducen el glutation oxidado a glutation reducido proceso catalizado por la enzima glutation reductasa, una flavoproteina con FAD. A su vez el glutation reducido retira el peroxido de hidrogeno, [H.sub.2][O.sub.2], del eritrocito en una reaccion catalizada por la glutation peroxidasa. Esta reaccion es importante, porque el [H.sub.2][O.sub.2] puede disminuir la esperanza de vida de los eritrocitos al incrementar la velocidad de la oxidacion de la hemoglobina a metahemoglobina (2) (Figura 1).

La deficiencia de G6PD provoca dano oxidativo irreversible y muerte celular (3). La vida media de 60 dias de la enzima refleja paso a paso la edad de los globulos rojos. Asi, a mayor edad, la actividad de algunas enzimas disminuye, pues los eritrocitos son incapaces de sintetizar nuevas moleculas proteicas. Por este motivo, los reticulocitos tienen una actividad enzimatica cinco veces mayor que la de los globulos rojos senescentes (4) y deben ser separados antes de efectuar la determinacion de la actividad de la enzima.

Estructura. La enzima glucosa-6-fosfato deshidrogenasa (E.C. 1.1.1.49; D-glucosa-6-fosfato: NADP oxidoreductasa) (5) esta presente en todas las celulas. En los eritrocitos se encuentra en sus formas dimerica y tetramerica. El monomero tiene un peso molecular de 59,256 daltons y consta de 515 aminoacidos. La actividad catalitica solo se inicia cuando se establece una asociacion en estado de equilibrio entre las formas dimerica y tetramerica (6). Tal asociacion requiere de la presencia de NADP, ligado fuertemente por la enzima (7,8), lo que hace que NADP desempene un papel dual, tanto de componente estructural como de coenzima9-11. En 1967, Luzzatto12 postulo por lo menos dos sitios de union del NADP a la enzima, con un estado de transicion de baja a alta afinidad cuando aumenta la concentracion de NADP, lo que significa que existen dos posibles formas de la enzima, de acuerdo con su afinidad por el NADP. Estas condiciones podrian variar en caso de aparicion de un inhibidor competitivo del NADPH. La baja constante de disociacion para el NADPH sugiere que la reaccion puede ser un controlador muy eficiente, a manera de retro-alimentador, que regularia la actividad enzimatica. Estos hechos permitieron establecer que la relacion de concentracion NADP/ NADPH es un mecanismo regulador de la actividad de la G6PD y por tanto de la via hexosa monofosfato (HMP) en el hematie.

Funcion. La importancia de la G6PD radica en la trascendencia de los procesos celulares en los que participa, a saber: Genesis de NADPH, efectuada a partir de los dos primeros pasos de la via hexosa monofosfato. El NADPH participa en la biosintesis reductora del colesterol y de los acidos grasos, asi como tambien en la sintesis del oxido nitrico (NO). Por otra parte se requiere para la actividad de la metahemoglobina reductasa y para el mantenimiento del nivel de glutation reducido (GSH). NADPH y GSH son los responsables del potencial redox efectivo para proteger del estres oxidativo tanto a los grupos sulfhidrilo de la membrana celular, como a las enzimas y a la hemoglobina que compromete la supervivencia del eritrocito (3).

Otras funciones, que muestran la trascendencia de esta enzima en la vida celular son:

1. Regulacion de la actividad de la proteina KU, implicada en reparar el ADN tras el dano que causan las radiaciones. La intervencion de la G6PD se efectua a traves del ciclo de las pentosas y consiste en facilitar la union de KU -con residuos de cisteina reducidos- al ADN en proceso de reparacion (13).

2. Desarrollo temprano del embrion. Cuando hay una deficiencia severa de G6PD en los tejidos extraembrionarios, el desarrollo de la placenta se detiene y se produce la muerte del embrion (14).

3. Supervivencia del feto durante la transicion de la hemoglobina fetal a la forma adulta. Aqui la G6PD impide el dano oxidativo debido a la generacion de especies reactivas de oxigeno a partir de la hemoglobina adulta (14,15).

4. Fagocitosis en celulas blancas. La deficiencia severa de esta enzima provoca una reduccion de la generacion de NADPH, lo que trae como resultado una disminucion de la produccion de [H.sub.2][O.sub.2], y por tanto la actividad microbicida del neutrofilo esta afectada, y asi mismo la respuesta inflamatoria (16). Aunque las caracteristicas clinicas de la deficiencia severa son semejantes a las de la enfermedad granulomatosa cronica (EGC), su aparicion ocurre, a diferencia de esta ultima, hacia las etapas de vida mas avanzadas (17,18). La EGC constituye un modelo fundamental para investigar la composicion y la activacion del sistema microbicida de las celulas fagociticas, en especial de los neutrofilos. Esta entidad se debe a un defecto profundo en la explosion respiratoria que acompana a la fagocitosis de todas las celulas mieloides (neutrofilos, eosinofilos, monocitos, macrofagos). La explosion respiratoria genera la conversion catalitica del oxigeno molecular en el anion superoxido que da lugar a la formacion de [H.sub.2][O.sub.2], de acido hipocloroso y de radicales hidroxilo. Estos derivados del oxigeno juegan un importante papel en la reaccion microbicida contra bacterias y hongos (19,20).

5. Modulacion del factor de crecimiento endotelial vascular que regula la angiogenesis. El NADPH se utiliza como cofactor de la oxido nitrico sintetasa endotelial (eNOS). Asi, el oxido nitrico requerido para la modulacion del crecimiento y la migracion endotelial durante el crecimiento vascular, se mantiene en un nivel adecuado (17).

6. La mayoria de los genes capaces de reducir el riesgo contra ciertas infecciones como la malaria se expresan en el globulo rojo, lo que se considera como un mecanismo genetico y/o evolutivo de defensa, como en el caso de los genes que expresan la G6PD (21).

Deficiencia. La deficiencia de G6PD aun prevalece como el mas comun de todos los defectos enzimaticos heredables (22,23) y clinicamente significativos, no solo en el campo de la hematologia, sino tambien de la biologia humana (24) y se caracteriza por una amplia heterogeneidad bioquimica y genetica.

La deficiencia de la G6PD ha sido el prototipo, dentro de las anemias hemoliticas, debida a una enzimopatia como anormalidad primaria del eritrocito. De igual manera es un ejemplo de anemia hemolitica debido a una interaccion entre causas extracelulares e intracelulares, ya que la hemolisis en la mayoria de los casos es disparada por agentes exogenos (25).

La hemolisis de los hematies deficientes ocurre como consecuencia del aumento en la susceptibilidad al dano oxidativo, debido a la incapacidad de las celulas para reducir de forma normal el NADP a NADPH. En presencia de agentes oxidantes, la produccion de NADPH a traves de la via HMP se estimula multiples veces, de modo que los niveles de NADPH y de GSH se mantienen estables. Estos eventos obedecen a la sobre-expresion de G6PD (26). El mecanismo exacto en el incremento de la sensibilidad al dano oxidativo, facilitador de la hemolisis, no es claro aun. Sin embargo, existe un significativo volumen de informacion sobre el favismo, mayor que el disponible acerca de los diferentes medicamentos que lo pueden producir. En las habas existen sustancias como la devicina y el isouramil, que producen oxidacion irreversible de GSH y de otros grupos de proteinas unidas por grupos -SH. Esto favorece en el hematie no solo un desequilibrio electrolitico, sino tambien la union por entrecruzamiento de las membranas y microvesiculizacion, eventos acompanados por un aumento en la concentracion de calcio en el eritrocit[O.sub.2]7.

La deficiencia de G6PD se produce por diversos mecanismos geneticos como deleciones, mutaciones puntuales y sustituciones que afectan la transcripcion, procesamiento o estructura primaria de la enzima, lo que funcionalmente lleva a una disminucion de la actividad enzimatica o perdida de afinidad por el sustrato. Hay otros factores que influyen sobre la actividad de la enzima. Asi, en un estudio cuyo objetivo era determinar la posible relacion entre la actividad de la enzima G6PD y la hipoxia, se encontro que la hipoxia favorecia una disminucion en su actividad (27,28).

Variantes. La deficiencia franca de G6PD se identifico inicialmente a mediados del siglo pasado, en norteamericanos de raza negra, en el curso de investigaciones llevadas a cabo sobre el efecto hemolitico de la primaquina (19). En la actualidad el medicamento continua como agente causal de la deficiencia en soldados iraquies con malaria (29).

Desde mediados del siglo pasado, se acepto que el defecto metabolico primario en sujetos susceptibles a la hemolisis secundaria a medicamentos o al consumo de habas (Vicia faba), corresponde a una baja actividad de la G6PD en los eritrocitos (30). Si bien esta claramente definida la asociacion entre la deficiencia de G6PD y la anemia hemolitica no inmune y no esferocitica (22), tambien es evidente su correlacion con la hemolisis debida a medicamentos, a alimentos y a otros eventos como procesos infecciosos, situacion que Vulliamy et al. (31) han destacado como el mas importante desencadenante de hemolisis. Hacia 1958, Gross et al. (30), por un lado y Szeinberg et al. (22) por otro, determinaron que la deficiencia enzimatica tenia una base hereditaria y sugirieron que estaba ligada al sexo.

La caracterizacion bioquimica ha permitido identificar no menos de 442 variantes de la deficiencia de la enzima. Alrededor de 299 fueron descritas mediante metodos utilizados por el grupo experto de la Organizacion Mundial de la Salud (OMS). Por otra parte, se documentaron 60 mutaciones o combinaciones, todas de naturaleza puntual, si se tiene en cuenta que la deficiencia total es incompatible con la vida (23).

De acuerdo con su nivel de actividad las variantes de la enzima se agruparon en cinco clases (23), que son:

Clase 1: Deficiencia de la enzima con anemia cronica hemolitica no esferocitica (CNSHA).

Clase 2: Deficiencia enzimatica severa (menos de 10%, por ejemplo, la forma mediterranea).

Clase 3: Deficiencia enzimatica moderada (10%-60%, por ejemplo, la forma africana).

Clase 4: Deficiencia enzimatica leve o ausente (60%-100%).

Clase 5: Actividad enzimatica por encima de lo normal.

La variante clase 1 es una forma rara y severa, asociada con la anemia hemolitica no esferocitica cronica. De aparicion esporadica, sus casos se consideran unicos (31). En regiones como los continentes africano y asiatico y la cuenca mediterranea existe una alta frecuencia de las diferentes variantes de la deficiencia enzimatica, mientras que en China y Japon la frecuencia es baja (23).

En las poblaciones mediterraneas la deficiencia enzimatica es mucho mas severa y frecuente que en la poblacion de raza negra norteamericana (32), donde el defecto se identifico en los hematies. En contraste, este fue hallado en varios tipos celulares diversos, obtenidos en individuos sensibles italianos y de raza judia (33). Con respecto a la frecuencia de la deficiencia severa, es notoria la variacion entre las distintas poblaciones. Asi, entre los americanos de raza negra, la frecuencia del gen de la deficiencia enzimatica es de 0.10% a 0.11% (34) con 15% de actividad enzimatica respecto a la normal35. Como ejemplo de una frecuencia elevada de la deficiencia, se puede citar a los judios kurdos en quienes alcanza, en su forma mediterranea, un valor igual a 0.7% (36). La forma mediterranea es una variante cuya frecuencia de polimorfismo tiene una actividad menor a 10%. En ella, la mutacion se presenta en el aminoacido 188, con sustitucion de fenilalanina (Phe) por serina (Ser) (37).

En Arabia Saudita la variante mas frecuente es la mediterranea, con frecuencias que oscilan entre 0% y 0.4% en hombres y 0% y 0.2% en mujeres. Es posible que la alta prevalencia en mujeres obedezca a una disomia uniparental o bien, a la alta consanguinidad existente o a que el cromosoma X que contiene el gen normal sea el que se inactiva durante la impronta genetica (38).

En latinoamerica se han descrito algunas variantes de la enzima. En Mexico por ejemplo, se identificaron 18, que son tambien de comun aparicion en otras regiones como el continente africano, el sur de Europa y el sudeste asiatico (39). Mientras que en Mexico la frecuencia de la deficiencia estuvo entre 0.4% y 4.1%, en Cuba fue 4.9% con una prevalencia de la variante A-, y 7% para la variante A+40. Para paises como Colombia, la frecuencia calculada por la OMS para las variantes fenotipicamente asociadas con la deficiencia severa (clase 2, con actividad menor de 10%) es entre 3% y 7% (23). Sin embargo, en 103 individuos de sexo masculino, donantes del Banco de Sangre de la Cruz Roja Colombiana y en apariencia sanos, se hallo una frecuencia de actividad subnormal (<60%) de aproximadamente 19.4%. Este estudio se hizo entre junio y octubre de 2003, mediante la aplicacion de la tecnica cualitativa de Beutler E (Palomino F. 2003. Universidad Nacional de Colombia. Comunicacion personal).

Otra clasificacion compara la movilidad electroforetica de las diversas variantes con la enzima normal B, siendo la variante A-, presente en individuos de raza negra con baja actividad enzimatica, mas rapida en un pH alcalino que la enzima normal, en contraste, la variante de los sujetos deficientes mediterraneos se mueve a una velocidad normal (37).

Otra variante comun, A+, tiene una actividad normal y se encuentra en mas o menos 20% de los norteamericanos de raza negra. Esta variante es electroforeticamente mas rapida que la B, hecho explicable si se tiene en cuenta que la sustitucion de asp (aminoacido neutro) por asn (aminoacido acido) en la posicion 126 modifica la carga electrica de la enzima, lo que se refleja en una movilidad electroforetica mas rapida (37).

La variante A- se encuentra en cerca de 11% de la poblacion negra norteamericana. No obstante, su frecuencia es mayor en la poblacion del Africa negra subsahariana. La actividad enzimatica de esta variante corresponde a 5% y 15% de la normal, disminucion debida a la presencia de dos sustituciones, no de una como ocurre en la variante A+. Una de esas sustituciones es identica a la que aparece en la variante A+ y la otra, unica para esta variante, obedece al cambio de val por met en la posicion 68 (41).

Genetica. A la enzima G6PD la codifica un gen presente en la region terminal del brazo largo del cromosoma X, (Xq28), menos de 2 centi-Morgan al gen del factor VIII. En los hombres, la condicion hereditaria ligada a X determina su caracter hemicigotico, lo que significa que hay un solo alelo, debido a la ausencia del locus homologo. Tambien hay mujeres homocigotas en poblaciones cuya frecuencia de la deficiencia de G6PD es alta. Las mujeres heterocigotas son portadoras aunque pueden desarrollar ataques hemoliticos. El gen de la G6PD se ha ubicado en la parte distal del brazo largo, tiene 18 Kb de largo y consta de 13 exones (42) (Cuadro 1).

La region del gen que codifica para la proteina comprende 12 segmentos, con un promedio de tamano entre 12 y 236 bp y un intron presente en la region no traductora 5'. En muchas lineas celulares, el extremo mayor 5' del ARNm de la G6PD se localiza a una distancia de 177 bp <<upstream>> del codon de iniciacion de la transcripcion (43,44). Aunque las mutaciones se extienden a lo largo de la region codificadora del gen, existen unas pocas (4 de 56) que dan origen a la forma mas severa de deficiencia de la enzima, esto es, la que se encuentra asociada con CNSHA (clase 1) en los 160 aminoacidos del extremo N-terminal. No obstante, no hay ninguna que cause formas moderadas de deficiencia (clases 2 y 3) en los 48 aminoacidos del extremo C-terminal. Muchas variantes en esta region exhiben movilidades electroforeticas anormales y son particularmente inestables cuando la concentracion de NADP es baja. Esto se debe a que esta region codifica para el dominio de union al NADP (45).

En la variante A- se presenta una sustitucion identica a la A+, aunque hay una segunda sustitucion en el nucleotido 202 G --> A del exon 4, lo que provoca el cambio val por met, acompanado por inestabilidad de la enzima in vivo. Asi, la diferencia entre las formas A y B corresponde al aminoacido que ocupa la posicion 126, presumiblemente como resultado de un empalme alternativo o <<splicing>> de considerable heterogeneidad entre los diferentes cADNs de la G6PD (41).

Manifestaciones clinicas. Casi todas las personas que cursan con la deficiencia de G6PD son usualmente asintomaticas y solo se manifiesta la enfermedad cuando ingieren drogas o quimicos que desencadenan la hemolisis masiva intravascular. La expresion clinica entonces resulta de la interaccion de las propiedades moleculares de cada variante de G6PD y de factores exogenos. Se han descrito diferentes sindromes clinicos asociados con la deficiencia de esta enzima que incluyen:

Hemolisis inducida por farmacos. Clasicamente, luego de la ingestion de ciertos agentes como sulfamidas, antipireticos, nitrofuranos y medicamentos antimalaricos, como la primaquina y cloroquina, el paciente desarrolla fiebre, orina de color negro, ictericia y anemia. La necrosis tubular aguda puede complicar el episodio hemolitico severo, sobre todo en las enfermedades subyacentes del higado, como hepatitis. El mantenimiento del flujo renal adecuado de la sangre, por diuresis alcalina forzada, puede prevenir esta complicacion. En estos casos con flujo renal comprometido de la sangre segun lo evidenciado por la salida baja de la orina, la transfusion es lo ideal con el proposito de eliminar las celulas rojas danadas que bloquean la microcirculacion y puede tambien evitar la complicacion renal. En algunos pacientes, la coagulacion intravascular diseminada (DIC) puede complicar la hemolisis intravascular masiva y necesitar el tratamiento apropiado (45).

Hemolisis inducida por infeccion. La infeccion es quiza la causa mas comun en quienes sufren deficiencia de G6PD. El mecanismo de hemolisis inducida por infecciones no es bien conocido; una explicacion puede ser que la generacion de [H.sub.2][O.sub.2] por los neutrofilos polimorfonucleares puede provocar una disminucion en la cantidad de glutation reducido, cuya funcion es eliminar del globulo rojo la acumulacion de metabolitos que oxidan a los grupos sulfhidrilos formados por el estres oxidativo, por lo que disminuye la capacidad protectora de la celula. Por otra parte, la activacion de los neutrofilos interviene directamente en la peroxidacion de los lipidos de la membrana y provoca de forma directa la destruccion de la celula (46). La severidad y las consecuencias clinicas estan influidas por muchos factores que incluyen la administracion simultanea de medicamentos oxidantes, los niveles de hemoglobina previos, la funcion hepatica y la edad (47).

Favismo. Se reconoce desde la antiguedad; los pacientes presentan un cuadro clinico similar al inducido por farmacos, que se desencadena dentro de las 24 y 48 horas siguientes a la ingesta de habas. Se caracteriza por la presencia de un cuadro de hemolisis aguda luego de ingerir habas, sin embargo, no todos los individuos con deficiencia de G6PD presentan hemolisis cuando comen habas (48).

Los sintomas del favismo se desarrollan pocas horas despues de la ingestion. Los mas comunes son nauseas, vomitos, malestar y vertigo. A estos sintomas les sigue una hemolisis aguda donde, a menudo, el recuento de eritrocitos cae por debajo de 1.0 x [10.sup.12]/l. En la mayoria de los globulos rojos aparecen cuerpos de Heinz. Estan presentes la hemoglobinemia y la hemoglobinuria. Los sintomas por lo general cesan luego de 2 a 6 dias (49,50).

Anemia hemolitica cronica no esferocitica. Las variantes de clase I, se caracterizan por este hallazgo, debido al grado tan severo de deficiencia enzimatica. La hemolisis es solo parcialmente intravascular y se puede acompanar de calculos biliares y esplenomegalia. Sin embargo, existe una variabilidad en las manifestaciones asociadas con este tipo de anemia cronica (47,50,51). En la literatura hay informes de casos donde describen anemia hemolitica cronica y una causa, aunque rara, es la deficiencia de G6PD (52).

Preeclampsia. Entidad al parecer asociada en forma parcial con una peroxidacion lipidica de la membrana plasmatica del sincitiotrofoblasto. De ahi que las mujeres con alguna alteracion en la actividad de la G6PD pueden tener serias dificultades en la reduccion del GSSG a GSH y asi presentar alteraciones en la defensa antioxidante (53).

De otro lado es interesante senalar como la deficiencia de G6PD (variante A-) se asocia con aumento en la resistencia a la infeccion por Plasmodium falciparum en la region subsahariana de Africa. Esto indica que hay una fuerte respuesta adaptativa frente a esta invasion del hematie (54).

CONCLUSIONES

Si se tiene en cuenta que Colombia ha sido asiento de multiples migraciones provenientes de Africa y Europa, es necesario llevar a cabo estudios que evaluen el rango de actividad de la G6PD, asi como sus posibles variantes en esta region. El conocimiento obtenido podra complementar los hallazgos efectuados en otras latitudes y por tanto derivara en un mejor diagnostico, enfoque y tratamiento de las enfermedades asociadas con la hemolisis y la oxidacion, como anemia, diabetes, hipertension arterial y cancer entre otras. Por otro lado, este conocimiento se podra aplicar en el area de la fisiologia del ejercicio, donde, a causa del estres oxidativo, pueden ocurrir desde hemolisis hasta la muerte subita del deportista.

En vista de los crecientes hallazgos sobre la participacion en ciertas entidades patologicas, asi como en la comprension de procesos evolutivos de la especie humana, la determinacion de la actividad de esta enzima es una prioridad para poblaciones como la colombiana donde solo se tienen datos extrapolados por la OMS.

En distintas partes del mundo surgen diferentes tecnicas que intentan favorecer la determinacion rapida con alta sensibilidad y especificidad de la actividad de esta enzima (55,56), sin embargo, para Colombia solo se han hecho algunas determinaciones de grupo pequenos (datos sin publicar) pero hay grupos de trabajo en ciertas universidades, como la Universidad del Rosario en Bogota, que pronto empezaran a dar resultados sobre la prevalencia de esta alteracion en el metabolismo de tipo hereditario.

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(27.) Turrini F, Naitana A, Mannuzzu L, Pescarmona G, Arese P. Increased red cell calcium adenosine triphosphatase, and altered membrane proteins during Fava bean hemolysis in glucose-6-phosphate dehydrogenase-deficient (Mediterranean variants) individuals. Blood 1985; 66: 302-305.

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JAVIER FERNANDO BONILLA, M.D., M.SC. (1), MAGDA CAROLINA SANCHEZ, LIC. QUIM. (2), LILIAN CHUAIRE, M.SC. (3)

(1.) Profesor Asistente, Facultad de Medicina y Facultad de Rehabilitacion, Universidad del Rosario, Bogota, Colombia. e-mail: jfbonill@urosario.edu.co

(2.) Profesora Asistente, Facultad de Medicina, Universidad del Rosario, Bogota, Colombia. e-mail: mcsanche@urosario.edu.co

(3.) Profesora Principal, Facultad de Medicina, Universidad del Rosario, Bogota, Colombia. e-mail: lchuaire@urosario.edu.co

Recibido para publicacion julio 12, 2005 Aceptado para publicacion enero 4, 2007

Glucose-6-phosphate dehydrogenase (G6PD). Response of the human erythrocyte and another cells to the decrease in their activity

JAVIER FERNANDO BONILLA, M.D., M.SC. (1), MAGDA CAROLINA SANCHEZ, LIC. QUIM. (2), LILIAN CHUAIRE, M.SC. (3)

SUMMARY

Glucose-6-phosphate dehydrogenase is the first enzyme in the pentose phosphate pathway and the main intracellular source of reduced nicotidamineadenine nucleotidephosphate (NADPH), involved in diverse physiological processes such as antioxidant defense, (for instance in the erythrocyte) endothelial growth modulation, erithropoyesis, vascularization and phagocitosis. G6PDH deficiency is the most common X-chromosome-linked enzymopathy in human beings. Although it is present in any type cell, its absolute deficiency is incompatible with life. According to WHO, 400 million people are affected by G6PD deficiency in the world but in Colombia, the severe form prevalence is about 3% to 7%. There are no data related to slight and moderate alterations, that also have clinical effects. This paper reviews some G6PD biomolecular aspects, its classification according to activity and electrophoretic mobility, as well as some main clinical aspects related to its activity alteration.

Keywords: Erythrocyte; Physiology; Genetics; Epidemiology; Glucose-6-phosphate dehydrogenase deficiency; Congenital hemolytic anemia.

Glucosa-6-fosfato deshidrogenasa (G6PD). Respuesta de los hematies y otras celulas humanas a la disminucion en su Actividad

RESUMEN

La glucosa-6-fosfato deshidrogenasa (G6PD) es la primera enzima de la via pentosa fosfato y la principal fuente intracelular de nicotidamina adenina dinucleotido fosfato reducido (NADPH), compuesto comprometido en diversos procesos fisiologicos, por ejemplo defensa antioxidante (sobre todo celulas como los eritrocitos), modulacion del crecimiento endotelial, eritropoyesis, vascularizacion y fagocitosis. La deficiencia de G6PD es la enzimopatia ligada al cromosoma X mas comun en el ser humano. Si bien se puede presentar en cualquier tipo de celula, su carencia absoluta es incompatible con la vida. Segun la OMS, en el mundo hay mas de 400 millones de personas afectadas por la deficiencia de la enzima, y para Colombia calculan una prevalencia de la deficiencia severa entre 3% y 7%, pero no se conocen los datos relativos a las alteraciones leves y moderadas, que tambien tienen efectos clinicos. El presente articulo revisa los aspectos biomoleculares mas importantes de la enzima, su clasificacion de acuerdo con la actividad y la movilidad electroforetica, y tambien se mencionan algunos aspectos clinicos relacionados con la alteracion de su actividad.

Palabras clave: Ultraestructura; Fisiologia; Genetica; Epidemiologia; Deficiencia de glucosafosfato deshidrogenasa; Eritrocitos; Anemia hemolitica congenita.

All living organisms, be yeasts or protozoa, plants or animals express the glucose-6-phosphate dehydrogenase enzyme (G6PD)(1).

Though G6PD is found in the cytoplasm of all mammals cells, its deficiency is more evident in the red blood cells probably because these cells live without nucleus for a long time and because they contain proteases that degrade the mutant enzyme in major degree than other proteases in other tissues (1).

Since the erythrocyte is a cell that transports oxygen by excellence, its mechanisms of defense against the oxidative stress make part of the maintenance of its circulating activity. These defense mechanisms depend in a great part on the metabolic supply of the reduced form of NADP (NADPH + H+). Due to the particular metabolic characteristics of these cells, only the two first reactions of the pentose via (also called hexose monophosphate) have the capacity to generate NADPH + H. These are first the conversion of the glucose-6-phosphate into phospho gluconic acid-6 and second the conversion of these intermediate one into ribulose-5-phosphate with CO2 detachment. These two reactions are sequential and in both the NADP is reduced. While the first one is catalyzed by the G6PD enzyme, the second one it is by the 6-phosphogluconate dehydrogenase.

Through the production of NADPH the erythrocytes reduce the oxidative glutathione to reduced glutathione which process is catalyzed by the glutathione reductase enzyme which is a flavoprotein with FAD (flavin adenin dinucleotic). At the same time the reduced glutathione retires the [H.sub.2][O.sub.2], from the erythrocyte in a reaction catalyzed by the peroxidase glutathione. This reaction is important because the [H.sub.2][O.sub.2] might reduce the life expectancy of the erythrocytes for the increase in the speed of hemoglobin oxidation to metahemoglobin (2) (Figure 1).

[FIGURE 1 OMITTED]

G6PD deficiency produces irreversible oxidative damage and cell death (3). The average life of 60 days of the enzyme reflects stepwise the age of the red blood cells. This way to major age the activity of some enzymes decreases since the erythrocytes are unable to synthesize new protein molecules. For this reason, the reticulocytes have an enzymatic activity five times major than that of the senescent (4) red blood cells and they must be separated before determining the enzyme activity.

Structure. The Glucose-6-phosphate dehydrogenase (E.C. 1.1.1.49; D-glucose-6-phosphate: NADP oxido-reductase) (5 ) enzyme is present in all cells. In the erythrocytes it is found in its dimeric and tetrameric forms. The monomer has a molecular weight of 59,256 daltons and counts with 515 amino acids. The catalytic activity is only initiated when an association is established, in balance status, between the dimeric and tetrameric (6) forms.

Such an association requires the NADP presence, strongly tied by the enzyme (7,8) which makes that NADP performs a dual role not only as structural component but as a coenzyme (9-11). In 1967, Luzzatto (12) postulated at least two sites of union of the NADP to the enzyme, with a condition of transition from low to high affinity when NADP's concentration increases, which means that there exist two possible forms of the enzyme, according to its affinity by the NADP. These conditions might change in case of appearance of a competitive inhibitor of the NADPH. The low constant of dissociation for the NADPH suggests that the reaction can be a very efficient controller, acting as a feedback, which would regulate the enzymatic activity. These facts allowed to establish that the relation of NADP/NADPH concentration is a regulatory mechanism of the activity of the G6PD and therefore of the hexose monophosphate (HMP) in the red blood cells.

Function. The importance of the G6PD is found in the transcendence of the cellular processes in which it takes part, such as:

NADPH genesis effected from the first two steps of the hexose monophosphate. The NADPH takes part in the biosynthesis that reduces cholesterol and oil acids and also in the synthesis of the nitric oxide (NO). On the other hand it is needed for the activity of the metahemoglobin reductase and for the maintenance of the reduced level of glutathione (GSH). NADPH and GSH are responsible for the potential redox effective to protect from the oxidative stress the groups of sulfhydryl of the cellular membrane, as well as the enzymes and the hemoglobin that compromises erythrocyte survival (3).

Other functions that show the transcendence of this enzyme in the cell life are the following:

1. Regulation of the activity of the KU protein implied in repairing the DNA after the damage that radiations cause. The intervention of the G6PD is effected through the pentose cycle and consists of facilitating the union of KU -with reduced cysteine residues- to the DNA in repairing process (13).

2. Early development of the embryo. When there is a severe G6PD deficiency in the extra-embryonic tissues, the placenta development stops and the embryo dies (14).

3. Survival of the fetus during the transition of the fetal hemoglobin to the adult form. Here the G6PD prevents the oxidative damage due to the generation of species reactive to oxygen from the adult hemoglobin (14,15).

4. Phagocytosis in white cells. The severe deficiency of this enzyme results in a reduction of the NADPH generation, which results in a decrease in the production of hydrogen peroxide ([H.sub.2][O.sub.2]) and therefore the neutrophil microbicide activity is affected and likewise its inflammatory response (16). Though the clinical characteristics of the severe deficiency are similar to those of the granulomatosa chronic disease (EGC), its appearance happens, unlike the latter, during more advanced stages of life (17,18). The EGC constitutes a fundamental model to investigate the composition and the activation of the microbicide system of the phagocyte cells, especially of the neutrophil ones. This entity is caused by a deep defect in the respiratory explosion that accompanies the phagocytosis of all myeloid cells (neutrophil, eosinophile, monocyte, macrophage). The respiratory explosion generates the catalytic conversion of the molecular oxygen in the super oxide anion that leads to the hydrogen peroxide formation, the hypochlorous acid and the hydroxile radicals. These derivates from the oxygen play an important role in the microbicide reaction against bacteria and fungi (19,20).

5. Modulation of the vascular endothelial factor growth that regulates the angiogenesis. NADPH is used as a cofactor of the endothelial nitric oxide sintetase (eNOS). Therefore the nitric oxide required for the modulation of the growth and for the endothelial migration during the vascular growth is maintained in an adequate level (17.)

6. Most of the genes able to reduce the risk against certain infections as the malaria are expressed in the red blood cell, which is considered like a genetic and/or evolutionary mechanism of defense, as in the case of the genes that express the G6PD (21).

Deficiency. The G6PD deficiency still prevails as the most common of all the inherited enzymatic defects (22,23) and it is clinically significant not only in the hematological field but also in the human biology (24) and it is characterized by a large biochemical and genetic heterogeneity. The deficiency of G6PD has been the prototype within the hemolytic anemias due to an enzymopathy as a primary abnormality of the erythrocyte. Likewise it is an example of hemolytic anemia due to an interaction between extra cellular and intracellular causes, since hemolysis in most cases is triggered by exogen agents (25).

Hemolysis of deficient red blood cells occurs as a consequence of the increase in the susceptibility to the oxidative damage due to the incapacity of cells to reduce the NADP to NADPH in a normal way. In presence of oxidant agents the NADPH production through the HMP is stimulated multiple times so that the NADPH and GSH levels maintain stable. These events are due to the overexpression of G6PD (26). The exact mechanism involved in the increase of the sensibility to the oxidative damage, facilitator of hemolysis, is not clear yet. However, there exists a significant volume of information on the favism, which is higher than the available one about the different medicines that might produce it. Faba beans contain compounds such as divicine and isouramil that produce irreversible GSH oxidation as well as in other groups of proteins united by SH groups. This favors not only an electrolytic unbalance in the red blood cells, but also the union by the intercrossing of membranes and microvesiculization, events accompanied by an increase in the calcium concentration in erythrocyte (27).

The G6PD deficiency is produced by different genetic mechanisms such as deletions, precise mutations and substitutions that affect the transcription, process or primary enzyme structure, which functionally results in a decrease of the enzymatic activity or loss of affinity by the substrate. There are other factors that influence the activity of the enzyme. Thus, in a study whose objective was to determine the possible relation between the activity of the G6PD enzyme and the hypoxia, it was found that the hypoxia favored a decrease in its activity (27,28).

Variants. The true deficiency of G6PD was identified initially in the middle of the last century, in black race North americans during the investigations carried out over the hemolytic effect of the primaquine (19). At the present time such medicine continues being a causal agent of the deficiency in Iraqi soldiers with malaria (29).

From mid of the last century, it was accepted that the primary metabolic defect in individuals susceptible to the hemolysis secondary to medicines or to the faba bean consumption (Vicia faba), corresponds to a low activity of the G6PD in erythrocytes (30). Although the association between the deficiency of G6PD and the non-immune hemolytic and the non-spherocytic anemia (22) is clearly defined, also it is evident its correlation with the hemolysis due to medicines, food and to other events such as infection processes, situation outlined by Vulliamy et al. (31) as the most important cause of hemolysis. Towards 1958, Gross et al. (30), on one hand and Szeinberg et al. (22) on the other one, determined that the enzymatic deficiency had a hereditary base and suggested that it was bound to sex. The biochemical characterization allowed identifying not less than 442 variants of the deficiency of the enzyme. About 229 of them were described by methods used by the expert group of the World Health Organization (WHO). On the other hand 60 mutations or their combinations were documented, all of precise nature taking into account that the total deficiency is incompatible with life (23).

According to its activity level the enzyme variants were classified in five types (23), such as:

Class 1: Deficiency of the enzyme with chronic non-spherlocytic-hemolytic anemia (CNSHA).

Class 2: Severe Enzymatic deficiency (less than 10%, for example the mediterranean form).

Class 3: Moderate enzymatic deficiency (10%-60%, for example, the African form).

Class 4: Low or absent enzymatic deficiency (60%-100%).

Class 5: Enzymatic activity above normal rates.

The class 1 variant is a rare and severe one, associated with chronic non-spherocytic hemolytic anemia. Of sporadic appearance, their cases are considered unique (31). In regions such as the African and Asian continents and the Mediterranean river basin there exists a high frequency of the different variants of the enzymatic deficiency, whereas in China and Japan the frequency is low (23).

In the Mediterranean populations the enzymatic deficiency is much more severe and frequent than in the population of North American black race (32), where the defect was identified in the red blood cells.

In contrast, this one was found in several diverse cellular types, obtained in Italian sensible individuals and of Jewish race (33). With respect to the frequency of the severe deficiency, the variation between the different populations is notorious. Thus, among the black race Americans, the frequency of the gene of the enzymatic deficiency is from 0.10% to 0.11% (34) with 15% of enzymatic activity related to normal (35). As an example of an elevated frequency of the deficiency, we may mention the Kurd Jews where it reaches, in its Mediterranean form, an equal value to 0.70% (36). The Mediterranean form is a variant whose frequency of polymorphism has an activity lower than 10%. In it, the mutation appears in the 188 amino acid, with substitution of phenylalanine (Phe) by serine (Ser) (37).

In Saudi Arabia the most frequent variant is the Mediterranean one, with frequencies that oscillate between 0% and 0.4% in men and 0% and 0.2% in women. It is possible that the high prevalence in women obeys to a uniparental dysomia or, to the high existing consanguinity or to that the chromosome X containing the normal gene is the one that inactivates itself during the genetic imprints (38).

In Latin America some variants of the enzyme have been described. In Mexico for example, 18 were identified, which are also of common appearance in other regions like the African continent, the south of Europe and the Southeast Asia (39). Whereas in Mexico the frequency of the deficiency was between 0.4% and 4.1%, in Cuba it was 4.9% with a prevalence of the variant A-, and 7% for the A+ variant (40). For countries like Colombia, the frequency calculated by the WHO for the variants phenotypically associated with the severe deficiency (Type 2, with lower activity than 10%) is between 3% and 7% (23). Nevertheless, in a survey developed among 103 individuals of masculine sex, donors of the Blood Donation Point of the Colombian Red Cross and in appearance healthy, it was found a frequency of subnormal activity (<60%) of approximately 19.4%. This research was developed between June and October of 2003, by means of the application of the qualitative technique of Beutler E (Palomino F. 2003. Universidad Nacional de Colombia, Perssonal communication).

Another classification is realized by comparing the electrophoretic mobility of the different variants with the normal B enzyme, being the variant A- enzyme, present in individuals of black race with low enzymatic activity, faster in alkaline pH than the normal enzyme, in contrast, the variant of the Mediterranean deficient individual's moves at a normal speed (37). Another frequent variant, A+, has a normal activity and it is found in about 20% of the North Americans of black race. This variant is electro-phoretically faster than the B, fact that is understood taking into account that the substitution of asp (neutral amino acid) by asn (acid amino acid) in 126 positions modifies the enzyme electrical charge, which is reflected in a faster electrophoretic mobility (37).

The variant A- is found in near 11% of the North American black population. However, its frequency is greater in the African sub-saharan black population. The enzymatic activity of this variant corresponds to 5% and 15% of the normal one, this decrease is due to the presence of two substitutions, not of one as it happens in the A+ variant. One of these substitutions is similar to the one that appears in variant A+ and the other one, unique for this variant, obeys to the change of the val for met in the 68 position (41).

Genetics. The G6PD enzyme is codified by a gene that is present in the terminal region of the long arm of chromosome X, (Xq28), less than 2 centi-Morgan to the gene of factor VIII. In men, the hereditary condition linked to X determines its hemizygotic character, which means that there is a single one allele, due to the absence of locus homologous. There are also homozygotic women in populations where the frequency of the G6PD deficiency is high. Hemocygotic women are carriers although they might develop hemolytic attacks. The G6PD gen has been mapped in the distal part of the long arm, its length is 18 Kb and counts with 13 exons (42) (Table 1).

The region of the gene to be codified for the protein includes 12 segments, with an average size between 12 and 236 bp and one intron present in the non-translator region 5'. In many cellular lines, the greater end 5' of the G6PD mRNA is located at a distance of 177 bp <<upstream>> from the transcription initiation codon (43,44). Although the mutations extend throughout the coding region of the gene, there exist few (4 of 56) that give origin to the most severe form of deficiency of the enzyme, this is, the one that is associated with CNSHA (type 1) in the 160 amino acids of the N-terminal end. However, there is no one that causes moderate forms of deficiency (types 2 and 3) in the 48 amino acids of the C-terminal end. Many variants in this region exhibit abnormal electrophoretic mobilities and are particularly unstable when the NADP concentration is low. This is because this region codifies for the domain of the union to NADP (45).

In the A- variant a substitution identical to A+ is present, although there is a second substitution in the 202 G ---> A nucleotide of exon 4 which results in the change of val for met, accompanied by the instability of the enzyme in vivo. Therefore the difference between forms A and B corresponds to the amino acid that occupies the 126 position, probably as a result of an alternate join or <<splicing>> of considerable heterogeneity among the different cDNAs of the G6PD (41).

Clinical manifestations. Almost all the people who attend with the G6PD deficiency are usually asymptomatic and the disease is only pronounced when they ingest drugs or chemicals that trigger the intravascular massive hemolysis. The clinical expression then results from the interaction of the molecular properties of each G6PD variant and from exogenous factors. Different clinical syndromes associated with the deficiency of this enzyme have been described and they include:

Hemolysis induced by drugs. Classically, after ingestion of certain agents like sulfamides, antipyretics, nitrofurans and antimalarial medicines, like the primaquine and chloroquine, the patient develops fever, black color urine, ictericia and anemia. The acute tubular necrosis might complicate the severe hemolytic episode, mainly in the underlying diseases of the liver, like hepatitis. The maintenance of the adequate renal flow of the blood, by forced alkaline diuresis, might prevent this complication. In these cases with the committed renal flow of the blood as evidenced by the low flow of urine, the transfusion is the ideal in order to eliminate the damaged red cells that block the microcirculation and can also avoid the renal complication. In some patients, the disseminated intra-vascular coagulation (DIC) might complicate the massive intravascular hemolysis and might need the appropriate treatment (45).

Hemolysis induced by infection. The infection is perhaps the most common cause in patients with G6PD deficiency. The mechanism of hemolysis induced by infections is not well known yet; an explanation may be that the generation of hydrogen peroxide by the polymorphonuclear neutrophiles might cause a decrease in the quantity of reduced glutathione, whose function is to eliminate, from the red cell, the accumulated metabolites that oxidate the sulfhydryl groups formed by the oxidative stress and therefore decreasing the protective capacity of the cell. On the other hand, the activation of the neutrophiles takes part directly in the peroxidation of lipids of the membrane and causes directly the destruction of the cell (46). The severity and clinical consequences are influenced by many factors that include the simultaneous administration of oxidating medicine, the previous levels of hemoglobin, the hepatic function and the age (47).

Favism. It is recognized from the antiquity; patients present a clinical picture similar to the induced by drugs, that is triggered within the following 24 to 48 hours after the ingestion of faba beans. It is characterized by the presence of a picture of acute hemolysis after ingesting faba beans; however, not all individuals with G6PD deficiency present hemolysis when ingesting faba beans (48).

Favisms symptoms are developed a few hours after the ingestion. The most common are nausea, vomit, discomfort and dizziness. After these symptoms there occurs an acute hemolysis where frequently the recount of erythrocytes falls below 1.0 x 1012/l. In most red blood cells Heinz bodies appear. The hemoglobinemia and hemoglobinuria are present. Symptoms stopped generally after 2 to 6 days (49,50).

Nonspherocytic chronic hemolytic anemia. The variants of type I, are characterized by this finding, due to the so severe degree of enzymatic deficiency. The hemolysis is only partially intravascular and it is possible to be accompanied by biliary calculous and splenomegaly. Nevertheless, it exists a variability in the manifestations associated with this type of chronic anemia (47,50,51). At the same time there are reports on cases in literature where they describe to chronic hemolytic anemia and a cause, although rare, is the G6PD deficiency (52).

Preeclampsia. Entity apparently associated in a partial form with a lipidic peroxidation of the plasmatic membrane of the syncitiotrophoblast. For that reason, patients with some alteration in the G6PD activity might have serious difficulties in the reduction of the GSSG to GSH and thus present alterations in the antioxidant defense (53).

On the other hand, it is interesting to outline how the G6PD deficiency (variant A-) is associated with an increase in the resistance to the infection caused by Plasmodium falciparum in the sub-Sahara region of Africa. This indicates that there is a strong adaptation response in front of this red blood cells invasion (54).

CONCLUSIONS

Considering that Colombia has been seat of multiple migrations from Africa and Europe, it is necessary to develop surveys in order to evaluate the rank of activity of the G6PD, as well as its possible variants in this region. The obtained knowledge will be able to complement the findings collected in other latitudes and therefore it will derive in a better diagnosis, approach and treatment of the diseases associated with the hemolysis and the oxidation, like anemia, diabetes, arterial hypertension and cancer among others. On the other hand, this knowledge might be applied in the area of the physiology of the exercise, where, because of the oxidative stress, it could happen from hemolysis to the sudden death of the sportsman.

Considering the increasing findings on the participation in certain pathologies, as well as in the understanding of evolutionary processes of the human species, the determination of the activity of this enzyme is a priority for populations like the Colombian one where the are only some extrapolated data obtained by WHO.

In different parts from the world different techniques arise that intend to favor the fast determination with high sensitivity and specificity of the activity of this enzyme (55,56). However, In Colombia only some determinations have taken place in small groups (data without publishing), and there exist small work groups from certain universities like the Universidad del Rosario of Bogota's, who will soon give their results in relation to the prevalence of this alteration in the metabolism of hereditary type.

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JAVIER FERNANDO BONILLA, M.D., M.SC. (1), MAGDA CAROLINA SANCHEZ, LIC. QUIM. (2), LILIAN CHUAIRE, M.SC. (3)

(1.) Assistant Professor, Facultad de Medicina y Facultad de Rehabilitacion, Universidad del Rosario, Bogota, Colombia. e-mail: jfbonill@urosario.edu.co

(2.) Assistant Professor, Facultad de Medicina, Universidad del Rosario, Bogota, Colombia. e-mail: mcsanche@urosario.edu.co

(3.) Principal Professor, Facultad de Medicina, Universidad del Rosario, Bogota, Colombia. e-mail: lchuaire@urosario.edu.co

Recibido para publicacion julio 12, 2005 Aceptado para publicacion enero 4, 2007
Cuadro 1
Caracteristicas moleculares del gen G6PD

ADN        Localizacion                    Xq2.8
           Tamano del gen (en kilobases)   18.5
           Numero de exones                13
           Numero de intrones              12

ARNm       Tamano (en nucleotidos)         2269
Proteina   Numero de aminoacidos           515
           Peso molecular (en daltons)     59,265
           Subunidades por molecula de
           enzima activa                   2 o 4

Table 1
Molecular characteristics of the G6PD gene

DNA       Localization                    Xq2.8
          Gen size (in kilobasis)         18.5
          Number of exons                 13
          Number of introns               12
mRNA      Size (in nucleotides)           2269
Protein   Number of amino acids           515
          Molecular weight (in daltons)   59,265
          Subunits by molecule of
          Active enzyme                   2 or 4
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Author:Bonilla, Javier Fernando; Sanchez, Magda Carolina; Quim, Lic; Chuaire, Lilian
Publication:Colombia Medica
Date:Jan 1, 2007
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