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Electric field measurements reveal the pivotal role of Cofactor-Substrate interaction in dihydrofolate reductase catalysis

Adesina, Aduragbemi S., Swiderek, Katarzyna, Luk, Louis Y. P., Moliner, Vicent and Allemann, Rudolf K. 2020. Electric field measurements reveal the pivotal role of Cofactor-Substrate interaction in dihydrofolate reductase catalysis. ACS Catalysis 10 (14) , pp. 7907-7914. 10.1021/acscatal.0c01856

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Abstract

The contribution of ligand–ligand electrostatic interaction to transition state formation during enzyme catalysis has remained unexplored, even though electrostatic forces are known to play a major role in protein functions and have been investigated by the vibrational Stark effect (VSE). To monitor electrostatic changes along important steps during catalysis, we used a nitrile probe (T46C-CN) inserted proximal to the reaction center of three dihydrofolate reductases (DHFRs) with different biophysical properties, Escherichia coli DHFR (EcDHFR), its conformationally impaired variant (EcDHFR-S148P), and Geobacillus stearothermophilus DHFR (BsDHFR). Our combined experimental and computational approach revealed that the electric field projected by the substrate toward the probe negates those exerted by the cofactor when both are bound within the enzymes. This indicates that compared to previous models that focus exclusively on subdomain reorganization and protein–ligand contacts, ligand–ligand interactions are the key driving force to generate electrostatic environments conducive for catalysis.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Chemistry
Publisher: American Chemical Society
ISSN: 2155-5435
Funders: BBSRC
Date of First Compliant Deposit: 6 July 2020
Date of Acceptance: 19 June 2020
Last Modified: 13 Aug 2020 10:36
URI: http://orca.cf.ac.uk/id/eprint/133133

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