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|Title:||Importance of the β5−β6 Loop for the structure, catalytic efficiency and stability of carbapenem-hydrolyzing class D β-Lactamase subfamily OXA-143|
|Author:||Antunes, Victor U.|
Llontop, Edgar E.
Vasconcelos, Fernanda N. da Costa
de los Santos, Yossef Lopez
Oliveira, Ronaldo J.
Farah, Chuck S.
Favaro, Denize C.
|Abstract:||The class D beta-lactamase OXA-143 has been described as an efficient penicillinase, oxacillinase, and carbapenemase. The D224A variant, known as OXA-231, was described in 2012 as exhibiting less activity toward imipenem and increased oxacillinase activity. Additionally, the P227S mutation was reported as a case of convergent evolution for homologous enzymes. To investigate the impact of both mutations (D224A and P227S), we describe in this paper a deep investigation of the enzymatic activities of these three homologues. OXA-143(P227S) presented enhanced catalytic activity against ampicillin, oxacillins, aztreonam, and carbapenems. In addition, OXA-143(P227S) was the only member capable of hydrolyzing ceftazidime. These enhanced activities were due to a combination of a higher affinity (lower K-m) and a higher turnover number (higher k(cat)). We also determined the crystal structure of apo OXA-231. As expected, the structure of this variant is very similar to the published OXA-143 structure, except for the two M223 conformations and the absence of electron density for three solvent-exposed loop segments. Molecular dynamics calculations showed that both mutants experience higher flexibility compared to that of the wild-type form. Therefore, our results illustrate that D224A and P227S act as deleterious and positive mutations, respectively, within the evolutionary path of the OXA-143 subfamily toward a more efficient carbapenemase|
|Editor:||American Chemical Society|
|Appears in Collections:||IQ - Artigos e Outros Documentos|
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