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Structure-Function Studies of Hydrophobic Residues That Clamp a Basic Glutamate Side Chain during Catalysis by Triosephosphate Isomerase.

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Richard JP, Amyes TL, Malabanan MM, Zhai X, Kim KJ, Reinhardt CJ, Wierenga RK, Drake EJ, Gulick AM
OpenAccess logo Biochemistry 55 3036-47 (2016)
Related entries: 5i3f, 5i3g, 5i3h, 5i3i, 5i3k

Cited: 15 times
EuropePMC logo PMID: 27149328

Abstract

Kinetic parameters are reported for the reactions of whole substrates (kcat/Km, M(-1) s(-1)) (R)-glyceraldehyde 3-phosphate (GAP) and dihydroxyacetone phosphate (DHAP) and for the substrate pieces [(kcat/Km)E·HPi/Kd, M(-2) s(-1)] glycolaldehyde (GA) and phosphite dianion (HPi) catalyzed by the I172A/L232A mutant of triosephosphate isomerase from Trypanosoma brucei brucei (TbbTIM). A comparison with the corresponding parameters for wild-type, I172A, and L232A TbbTIM-catalyzed reactions shows that the effect of I172A and L232A mutations on ΔG(⧧) for the wild-type TbbTIM-catalyzed reactions of the substrate pieces is nearly the same as the effect of the same mutations on TbbTIM previously mutated at the second side chain. This provides strong evidence that mutation of the first hydrophobic side chain does not affect the functioning of the second side chain in catalysis of the reactions of the substrate pieces. By contrast, the effects of I172A and L232A mutations on ΔG(⧧) for wild-type TbbTIM-catalyzed reactions of the whole substrate are different from the effect of the same mutations on TbbTIM previously mutated at the second side chain. This is due to the change in the rate-determining step that determines the barrier to the isomerization reaction. X-ray crystal structures are reported for I172A, L232A, and I172A/L232A TIMs and for the complexes of these mutants to the intermediate analogue phosphoglycolate (PGA). The structures of the PGA complexes with wild-type and mutant enzymes are nearly superimposable, except that the space opened by replacement of the hydrophobic side chain is occupied by a water molecule that lies ∼3.5 Å from the basic side chain of Glu167. The new water at I172A mutant TbbTIM provides a simple rationalization for the increase in the activation barrier ΔG(⧧) observed for mutant enzyme-catalyzed reactions of the whole substrate and substrate pieces. By contrast, the new water at the L232A mutant does not predict the decrease in ΔG(⧧) observed for the mutant enzyme-catalyzed reactions of the substrate piece GA.

Articles - 5i3j mentioned but not cited (1)

  1. Structure-Function Studies of Hydrophobic Residues That Clamp a Basic Glutamate Side Chain during Catalysis by Triosephosphate Isomerase. Richard JP, Amyes TL, Malabanan MM, Zhai X, Kim KJ, Reinhardt CJ, Wierenga RK, Drake EJ, Gulick AM. Biochemistry 55 3036-3047 (2016)


Reviews citing this publication (4)

  1. Enzyme activation through the utilization of intrinsic dianion binding energy. Amyes TL, Malabanan MM, Zhai X, Reyes AC, Richard JP. Protein Eng Des Sel 30 157-165 (2017)
  2. A guide to the effects of a large portion of the residues of triosephosphate isomerase on catalysis, stability, druggability, and human disease. Olivares-Illana V, Riveros-Rosas H, Cabrera N, Tuena de Gómez-Puyou M, Pérez-Montfort R, Costas M, Gómez-Puyou A. Proteins 85 1190-1211 (2017)
  3. The role of ligand-gated conformational changes in enzyme catalysis. Moreira C, Calixto AR, Richard JP, Kamerlin SCL. Biochem Soc Trans 47 1449-1460 (2019)
  4. A reevaluation of the origin of the rate acceleration for enzyme-catalyzed hydride transfer. Reyes AC, Amyes TL, Richard JP. Org Biomol Chem 15 8856-8866 (2017)

Articles citing this publication (10)

  1. Protein Flexibility and Stiffness Enable Efficient Enzymatic Catalysis. Richard JP. J Am Chem Soc 141 3320-3331 (2019)
  2. Enzyme Architecture: Modeling the Operation of a Hydrophobic Clamp in Catalysis by Triosephosphate Isomerase. Kulkarni YS, Liao Q, Petrović D, Krüger DM, Strodel B, Amyes TL, Richard JP, Kamerlin SCL. J Am Chem Soc 139 10514-10525 (2017)
  3. Enzyme Architecture: Amino Acid Side-Chains That Function To Optimize the Basicity of the Active Site Glutamate of Triosephosphate Isomerase. Zhai X, Reinhardt CJ, Malabanan MM, Amyes TL, Richard JP. J Am Chem Soc 140 8277-8286 (2018)
  4. Uncovering the Role of Key Active-Site Side Chains in Catalysis: An Extended Brønsted Relationship for Substrate Deprotonation Catalyzed by Wild-Type and Variants of Triosephosphate Isomerase. Kulkarni YS, Amyes TL, Richard JP, Kamerlin SCL. J Am Chem Soc 141 16139-16150 (2019)
  5. Enzyme Architecture: Breaking Down the Catalytic Cage that Activates Orotidine 5'-Monophosphate Decarboxylase for Catalysis. Reyes AC, Plache DC, Koudelka AP, Amyes TL, Gerlt JA, Richard JP. J Am Chem Soc 140 17580-17590 (2018)
  6. Local interactions with the Glu166 base and the conformation of an active site loop play key roles in carbapenem hydrolysis by the KPC-2 β-lactamase. Furey IM, Mehta SC, Sankaran B, Hu L, Prasad BVV, Palzkill T. J Biol Chem 296 100799 (2021)
  7. Itavastatin and resveratrol increase triosephosphate isomerase protein in a newly identified variant of TPI deficiency. VanDemark AP, Hrizo SL, Eicher SL, Kowalski J, Myers TD, Pfeifer MR, Riley KN, Koeberl DD, Palladino MJ. Dis Model Mech 15 dmm049261 (2022)
  8. Kinetics and mechanism for enzyme-catalyzed reactions of substrate pieces. Cristobal JR, Richard JP. Methods Enzymol 685 95-126 (2023)
  9. The Role of Asn11 in Catalysis by Triosephosphate Isomerase. Hegazy R, Cordara G, Wierenga RK, Richard JP. Biochemistry 62 1794-1806 (2023)
  10. Triosephosphate Isomerase: The Crippling Effect of the P168A/I172A Substitution at the Heart of an Enzyme Active Site. Hegazy R, Richard JP. Biochemistry 62 2916-2927 (2023)


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