Estimation of binding constants for the substrate and activator of Rhodospirillum rubrum adenosine 5'-diphosphate-glucose pyrophosphorylase using affinity capillary electrophoresis

CHED 184

Taguhi Sogomonyan, Taguhi_sogomonyan@hotmail.com, Department of Biochemistry and Chemistry, California State University Los Angeles, 5151 State University Drive, Los Angeles, CA 90032, Christopher R. Meyer, cmeyer@fullerton.edu, Department of Chemistry and Biochemistry, California State University, Fullerton, 800 N. State College Blvd., Fullerton, CA 92834, and Frank A Gomez, cufomadu@yahoo.com, Department of Chemistry and Biochemistry, California State University, 5151 State University Drive, Los Angeles, CA 90032-8202.
ADP-glucose pyrophosphorylase (ADPGlc PPase) catalyzes the conversion of ATP and Glc-1-P to ADPGlc and pyrophosphate; ADPGlc serves as the glucosyl donor for enzymatic glucan production in bacteria and plants and is the rate-limiting allosteric enzyme in a pathway producing renewable and biodegradable carbon. The enzyme from Rhodospirillum rubrum (Rs.r.) is activated by pyruvate. Binding constants were determined for ATP and Glc-1-P with and without pyruvate, using affinity capillary electrophoresis (ACE). The capillary is injected with a sample containing ADPGlc PPase and standards. The sample is subjected to increasing concentrations of ATP, pyruvate, or Glc-1-P in the running buffer and electrophoresed. Examination of the change in the migration times of ADPGlc PPase, relative to those of the standards, as a function of the increasing concentration of ligand provides a binding constant. The method is being extended to determine binding constants for altered proteins that harbor mutations in putative binding sites.
 

Undergraduate Research Poster Session: Analytical Chemistry
2:30 PM-4:30 PM, Monday, 11 September 2006 Moscone Center -- Hall D, Poster

Division of Chemical Education

The 232nd ACS National Meeting, San Francisco, CA, September 10-14, 2006