Triosephosphate isomerase (TIM) is the perfectly evolved enzyme in glycolysis that functions to interconvert dihydroxyacetone phosphate (DHAP) and glyceraldehyde 3-phosphate (GAP). There are numerous studies on the catalytic function and structure of this particular enzyme, but it remains challenging to create enzymes with this high level of catalytic efficiency suggesting there are features of catalysis still not well understood. As enzymes are more than the few residues directly contacting the substrate, a great interest in better understanding the functions of enzymes has been to investigate not only the active site but also the significant residues surrounding the active site.
One of these key residues is Glu97. This residue is located close to the active site and is conserved throughout known TIM sequences. Due to these interesting features, Glu97 has been an important focus of research to further understand TIM’s structure and catalytic functionality. Although there are several studies to evaluate the role of Glu97 in several species, Glu97 in TIM from Saccharomyces cerevisiae (yTIM) has not been extensively studied. Herein, several mutations of Glu97 were made in yTIM and the effects of the mutations on the function and structure of the enzyme were evaluated. The present study shows that when Glu97 is mutated to Asp, Gln, Ala, and Gly in yTIM, the catalytic effects and changes in protein stability were small. The results from dimethyl suberimidate (DMS) cross-linking assays suggest that the Glu97 mutants maintained their dimeric structures. Based on the different functional effects between Glu97 mutants in yTIM compared to other TIM species, we propose that the variation of surrounding residues in the active site provides a wide range of deleterious effects and stresses the complexity of residue positioning in an enzyme structure.
|Commitee:||Bhandari, Deepali, Weers, Paul|
|School:||California State University, Long Beach|
|Department:||Chemistry and Biochemistry|
|School Location:||United States -- California|
|Source:||MAI 82/9(E), Masters Abstracts International|
|Keywords:||Saccharomyces cerevisiae, Triosephosphate isomerase, Non-active site residues, Enzyme structure|
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