Coordinated effects of distal mutations on environmentally coupled tunneling in dihydrofolate reductase

Lin Wang, Nina Goodey, Stephen J. Benkovic, Amnon Kohen

Research output: Contribution to journalArticle

135 Citations (Scopus)

Abstract

One of the most intriguing questions in modern enzymology is whether enzyme dynamics evolved to enhance the catalyzed chemical transformation. In this study, dihydrofolate reductase, a small monomeric protein that catalyzes a single C-H-C transfer, is used as a model system to address this question. Experimental and computational studies have proposed a dynamic network that includes two residues remote from the active site (G121 and M42). The current study compares the nature of the H-transfer step of the WT enzyme, two single mutants, and their double mutant. The contribution of quantum mechanical tunneling and enzyme dynamics to the H-transfer step was examined by determining intrinsic kinetic isotope effects, their temperature dependence, and activation parameters. Different patterns of environmentally coupled tunneling were found for these four enzymes. The findings indicate that the naturally evolved WT dihydrofolate reductase requires no donor-acceptor distance fluctuations (no gating). Both single mutations affect the rearrangement of the system before tunneling, so some gating is required, but the overall nature of the environmentally coupled tunneling appears similar to that of the WT enzyme. The double mutation, on the other hand, seems to cause a major change in the nature of H transfer, leading to poor reorganization and substantial gating. These findings support the suggestion that these distal residues synergistically affect the H transfer at the active site of the enzyme. This observation is in accordance with the notion that these remote residues are part of a dynamic network that is coupled to the catalyzed chemistry.

Original languageEnglish
Pages (from-to)15753-15758
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume103
Issue number43
DOIs
StatePublished - 24 Oct 2006

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Tetrahydrofolate Dehydrogenase
Mutation
Enzymes
Catalytic Domain
Isotopes
Temperature
Proteins

Keywords

  • Enzyme dynamics
  • Hydrogen transfer
  • Hydrogen tunneling
  • Kinetic isotope effect
  • Structure function

Cite this

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abstract = "One of the most intriguing questions in modern enzymology is whether enzyme dynamics evolved to enhance the catalyzed chemical transformation. In this study, dihydrofolate reductase, a small monomeric protein that catalyzes a single C-H-C transfer, is used as a model system to address this question. Experimental and computational studies have proposed a dynamic network that includes two residues remote from the active site (G121 and M42). The current study compares the nature of the H-transfer step of the WT enzyme, two single mutants, and their double mutant. The contribution of quantum mechanical tunneling and enzyme dynamics to the H-transfer step was examined by determining intrinsic kinetic isotope effects, their temperature dependence, and activation parameters. Different patterns of environmentally coupled tunneling were found for these four enzymes. The findings indicate that the naturally evolved WT dihydrofolate reductase requires no donor-acceptor distance fluctuations (no gating). Both single mutations affect the rearrangement of the system before tunneling, so some gating is required, but the overall nature of the environmentally coupled tunneling appears similar to that of the WT enzyme. The double mutation, on the other hand, seems to cause a major change in the nature of H transfer, leading to poor reorganization and substantial gating. These findings support the suggestion that these distal residues synergistically affect the H transfer at the active site of the enzyme. This observation is in accordance with the notion that these remote residues are part of a dynamic network that is coupled to the catalyzed chemistry.",
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Coordinated effects of distal mutations on environmentally coupled tunneling in dihydrofolate reductase. / Wang, Lin; Goodey, Nina; Benkovic, Stephen J.; Kohen, Amnon.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 103, No. 43, 24.10.2006, p. 15753-15758.

Research output: Contribution to journalArticle

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