Question

1. Below is the enzyme mechanism of an enzyme we will study next semester from a Biochemistry article (citation removed so you don't look up the answer). a. Which of the major catalytic mechanisms is this enzyme using? Label them on the mechanism. b. Which of the six classes of enzymes would this one belong to? How did you know?

Answer 1

Here the mechanism of Triose Phosphate Isomerase enzyme is taken.

Major Catalytic Mechanism

Isomerisation of Triose Phosphate Isomerase is carried through the acid base mediated mechanism. This reaction implies with three catalytic residues, which is individually in direct contact with the substrate. The active site of the enzyme attracts the GAP or DHAP substrate by electrostatic interactions among the positively charged Lys12, and the phosphate group which is negatively charged. The leads to the Substrate Stabilisation. Based on this mechanism, a general based catalyst is Glu165. It take away a proton from pro(R) position of carbon 1 of DHAP or C-2 proton of GAP. The basic character for taking away the proton will not owned by the Glutamate's Carboxyl group, and it needs His95. The general acid His95 is used for the stabilisation of the negatively charged C-2 carbonyl oxygen and also stabilise the planar endediole(ate) intermediate by donating a proton. This enededialo(ate) is also stabilised by Lys12 and Asn11.

During this Process, general based catalyst Glu165 plays a role of general acid, which gives a proton to the nearby C-2, whereas the general acid His95 possess a general base character, which attracts a proton from C-1 hydroxyl group. At last, the enzyme gets altered by the GAP Isomer formation. During that instant, histidine and glutamate get back to their primitive form. Observation of DHAP labeled as tritium shows that the intramolecular transfer of ³H label is only 6% granted for GAP product. In overall solvent, the hydrogen bonded with the Glu165 will be in equilibrium position. Methyl Glyoxal Synthase(MGS) which is obtained from the reaction mechanism of methy glyoxal is considered to be as same as triose phosphate isomerase. The two enzymes make use of DHAP to assemble the intermediate named enediol(ate) phosphate. The next step of catalysation will be in the MGS route shows that the phosphate gets ejected, and the enediol(ate) gets ruined. This results in the product formation of methylglyoxal, more or less GAP isomer is formed from the reprotonation.

About Enediol(ate) intermediate

Enediol(ate) acts as a Kinetic barrier, which associates with the attraction of the $\alpha$-proton from the substrate of carbon acid. The reason for this adverse effect is the larger pKa value of the substrate's C-1 proton, and acidity shortage. This thermodynamic barrier is overwhelmed by deprotonation of C1 and deprotonation of C2 carbonyl oxygen. This in turn gives way for equivalent free energies for each species, as a result the equilibrium constant urges near unity.

Energy profile for Triose Phosphate isomerase reaction catalysis

AG (KJ/MO1) 401 E-GAP E+ GAP 10+ E-DHAP E-enediol • EF DHAP Reaction Coordinate

2. This enzyme mechanism belongs to Triose Phosphate Isomerase.