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Role of the occluded conformation in bacterial dihydrofolate reductases

Behiry, Enas Mamdouh, Luk, Louis Yu Pan, Matthews, Stella M., Loveridge, Edric Joel and Allemann, Rudolf Konrad 2014. Role of the occluded conformation in bacterial dihydrofolate reductases. Biochemistry 53 (29) , pp. 4761-4768. 10.1021/bi500507v

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Abstract

Dihydrofolate reductase (DHFR) from Escherichia coli (EcDHFR) adopts two major conformations, closed and occluded, and movement between these two conformations is important for progression through the catalytic cycle. DHFR from the cold-adapted organism Moritella profunda (MpDHFR) on the other hand is unable to form the two hydrogen bonds that stabilize the occluded conformation in EcDHFR and so remains in a closed conformation during catalysis. EcDHFR-S148P and MpDHFR-P150S were examined to explore the influence of the occluded conformation on catalysis by DHFR. Destabilization of the occluded conformation did not affect hydride transfer but altered the affinity for the oxidized form of nicotinamide adenine dinucleotide phosphate (NADP+) and changed the rate-determining step of the catalytic cycle for EcDHFR-S148P. Even in the absence of an occluded conformation, MpDHFR follows a kinetic pathway similar to that of EcDHFR with product release being the rate-limiting step in the steady state at pH 7, suggesting that MpDHFR uses a different strategy to modify its affinity for NADP+. DHFRs from many organisms lack a hydrogen bond donor in the appropriate position and hence most likely do not form an occluded conformation. The link between conformational cycling between closed and occluded forms and progression through the catalytic cycle is specific to EcDHFR and not a general characteristic of prokaryotic DHFR catalysis.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Chemistry
Subjects: Q Science > QD Chemistry
Publisher: ACS Publications
ISSN: 0006-2960
Funders: BBSRC
Date of First Compliant Deposit: 30 March 2016
Last Modified: 18 Dec 2018 21:19
URI: http://orca.cf.ac.uk/id/eprint/63599

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Cited 1 time in Web of Science. View in Web of Science.

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