Triose Phosphate Isomerase Structure & Mechanism: Difference between revisions

<scene name='Christian_Krenk_Sandbox/Active_site/1'> Glu 165 acts as the base and grabs the C2 proton on glyceraldehyde-3-phosphate, while His 95 is H-bonded to the carbonyl oxygen and acts as the acid by protonating carbonyl oxygen.</scene>  The enediol intermediate is negatively charged, but is somewhat <scene name='Christian_Krenk_Sandbox/Lysine/1'>stabilized by the positively charged side chain of Lys 12.</scene> <ref name= "lodi">PMID:8130193</ref>  Mutation of Lys 12 to Arg increases Km by a factor of 22 and decreases Vmax by a factor of 180.<ref name="lodi" /> To convert the enediol intermediate to DHAP, C1 is protonated by Glu 165, with His 95 removing a proton from C2’s OH group.  As a result, the catalytic groups are back to their original states, and catalysis is complete.  With GAP as a substrate, Km for the reaction is .34 mM and Vmax is 7200 units/mg protein at 25 degrees C and pH 7.5.<ref name= "dab" />  

An interesting part of the enzyme is the <scene name='Christian_Krenk_Sandbox/Flexible_loop/1'>flexible loop</scene> that stabilizes the enediol-like transition state.  The flexible loop (residues 167-176)<ref>PMID:2204418</ref> closes over the active site like a hinged lid when substrate is bound, thus preventing phosphate from leaving.  A four-residue segment of the loop H-bonds with the phosphate group of the substrate.<ref name="book" />  Without the loop, the enediol intermediate would eliminate phosphate, with the end products being inorganic phosphate and toxic methylglyoxal.<ref name="book" />