Birgy André, Magnan Mélanie, Hobson Claire Amaris, Figliuzzi Matteo, Panigoni Karine, Codde Cyrielle, Tenaillon Olivier, Jacquier Hervé
IAME, UMR 1137, INSERM, Université de Paris Cite, 75014 Paris, France.
Service de Microbiologie, Hôpital Robert-Debré, AP-HP, 75019 Paris, France.
Antibiotics (Basel). 2022 May 13;11(5):652. doi: 10.3390/antibiotics11050652.
Due to their rapid evolution and their impact on healthcare, beta-lactamases, protein degrading beta-lactam antibiotics, are used as generic models of protein evolution. Therefore, we investigated the mutation effects in two distant beta-lactamases, TEM-1 and CTX-M-15. Interestingly, we found a site with a complex pattern of genetic interactions. Mutation G251W in TEM-1 inactivates the protein's function, just as the reciprocal mutation, W251G, does in CTX-M-15. The phylogenetic analysis revealed that mutation G has been entrenched in TEM-1's background: while rarely observed throughout the phylogeny, it is essential in TEM-1. Using a rescue experiment, in the TEM-1 G251W mutant, we identified sites that alleviate the deviation from G to W. While few of these mutations could potentially involve local interactions, most of them were found on distant residues in the 3D structure. Many well-known mutations that have an impact on protein stability, such as M182T, were recovered. Our results therefore suggest that entrenchment of an amino acid may rely on diffuse interactions among multiple sites, with a major impact on protein stability.
由于β-内酰胺酶（可降解β-内酰胺类抗生素的蛋白质）的快速进化及其对医疗保健的影响，它们被用作蛋白质进化的通用模型。因此，我们研究了两种远缘β-内酰胺酶TEM-1和CTX-M-15中的突变效应。有趣的是，我们发现了一个具有复杂遗传相互作用模式的位点。TEM-1中的G251W突变会使蛋白质功能失活，就像CTX-M-15中的反向突变W251G一样。系统发育分析表明，突变G已在TEM-1的背景中根深蒂固：虽然在整个系统发育过程中很少观察到，但它在TEM-1中是必不可少的。通过一个拯救实验，在TEM-1 G251W突变体中，我们确定了一些位点，这些位点减轻了从G到W的偏差。虽然这些突变中很少有可能涉及局部相互作用，但大多数是在三维结构中距离较远的残基上发现的。许多对蛋白质稳定性有影响的著名突变，如M182T，也被发现。因此，我们的结果表明，氨基酸的固定可能依赖于多个位点之间的分散相互作用，对蛋白质稳定性有重大影响。
