25-30 August 2019
Henry Ford Building
Europe/Berlin timezone

ROLE OF CONFORMATIONAL FLEXIBILITY IN THE FITNESS OF β-LACTAMASES ENZIMES

Not scheduled
4h
Harnack House and Henry Ford Building

Harnack House and Henry Ford Building

Board: 393
Poster Posters

Speaker

Ms Maria Agustina Rossi (IBR-CONICET)

Description

Alternate structural conformers are the foundation of “protein evolvability”, which refers to the aptitude of proteins to rapidly adopt new functions within existing folds or even adopt entirely new folds. The understanding of protein evolution depends on the ability to relate the impact of mutations on molecular traits to organismal fitness. Antibiotic resistance mediated by β-lactamases represents an evolutionary paradigm in which the organismal fitness depends on the catalytic efficiency of a single enzyme. Here, we analyze the evolution of CTX-M-14, a serine- β-lactamase into the clinical variant CTX-M-16 (D240G, V231A) with higher activity against ceftazidime, one of the bulkier cephalosporins, and cefotaxime, the natural substrate. CTX-M-27 (D240G) displayed a reduced catalytic efficiency against cefotaxime but it improves against ceftazidime and CTX-M-9 (V231A) showed no activity difference. Tthese expanded hydrolytic activities are generally related to an enlargement of the active site, but this is not the case for CTX-M. We hypothesized that this could be due to an increased flexibility.
To test this hypothesis, we studied the dynamics of CTX-M-14 and its natural variants by NMR. The V231A mutation affects the CSP of the residues located on the αβ domain and the D240G does it on the Ω-loop, an essential catalytic structure of SBLs. The ps-ns dynamics did not show any important difference between the enzymes. However, in H/D exchange experiments the double mutant displayed a much faster exchange, particularly than the wild-type enzyme, of the residues located on or near the active. These findings may suggest that, in the clinical environments, CTX-M-14 evolved to generate a variant with an increased flexibility that is able to bind and hydrolyze larger substrates.

Primary authors

Ms Maria Agustina Rossi (IBR-CONICET) Prof. Alejandro Jose Vila (IBR-CONICET)

Presentation Materials

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