Screening for mutant clones of creatine kinase. (Medical Science and Health Poster Session 09:00 AM-10:00 AM).
Creatine kinase (CK) is a 86 kDa homodimer that catalyzes the reversible synthesis of adenine triphosphate (ATP) from the hydrolysis of phosphocreatine to creatine. This reaction maintains an energy buffer in a variety of cells (muscle, brain, heart, and mitochondrial) and serves as a transportation system that moves energy around the cell. By studying the structure and function of CK, we can learn how structure dictates function, as well as information about CK on a molecular level, such as the discovery of second-site suppressors. The purpose of this research project is to develop a plate assay that will easily screen for the restored activity of an inactive mutant CK. The plate assay will occur on a petri plate in which E. coli cells (BL-21) are induced to secrete CK. Once secreted, the protein will react with chemicals in the media to create an insoluble, purple precipitate, which indicates active CK. This will allow the researcher to rapidly screen for new mutants or second site suppressors of already characterized mutants. An enhanced pET17 clone of the CK gene is currently being constructed to facilitate the secretion of CK by cloning in secretion tag sequences. This engineered construct will be used in the development of the plate assay.
JENNIFER M. DLWGOSH JDLWGOSH@WOOSTER.EDU, (DEAN FRAGA DFRAGA@WOOSTER.EDU), COLLEGE OF WOOSTER, C-1412 1189 BEALL AVE, WOOSTER OH 44691
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|Author:||Dlwgosh, Jennifer M.; Fraga, Dean|
|Publication:||The Ohio Journal of Science|
|Date:||Mar 1, 2003|
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