Synthesis of b-lactamases is one of the common mechanisms of bacterial resistance to b-lactam antibiotics including penicillins and cephalosporins. The widespread use of antibiotics results in appearance of numerous extended-spectrum b-lactamase variants or resistance to inhibitors. Mutations of 92 residues of TEM type were found. Several mutations are the key mutations that determine the extension of spectrum of substrates. However, roles of the most associated mutations, located far from active site, remain unknown. We have investigated the role of associated mutations in structure of b-lactamase TEM-72, which contain two key mutation (G238S, E240K) and two associated mutations (Q39K, M182T) by means of simulation of molecular dynamics. The key mutation lead to destabilization of the protein globule, characterized by increased mobility of amino acid residues at high temperature of modelling. Mutation M182T lead to stabilization protein, whereas mutation Q39K is destabilizing mutation. It seems that the last mutation serves for optimization of conformational mobility of b-lactamase and may influence on enzyme activity.
Sintez b-laktamaz iavliaetsia odnim iz naibolee obshchikh mekhanizmov rezistentnosti bakteriĭ k b-laktamnym antibiotikam, takim kak penitsilliny i tsefalosporiny. Shirokoe ispol'zovanie antibiotikov privelo k vozniknoveniiu mnozhestva variantov b-laktamaz s rasshirennym spektrom deĭstviia ili ustoĭchivym k ingibitoram. U b-laktamaz TEM tipa uzhe opisany mutatsii 92 aminokislotnykh ostatkov. Riad mutatsiĭ iavliaiutsia funktsional'nymi i otvechaiut za rasshirenie spektra substratnoĭ spetsifichnosti. Odnako rol' bol'shinstva tak nazyvaemykh soputstvuiushchikh mutatsiĭ, raspolozhennykh vdali ot aktivnogo tsentra, ostaetsia neizuchennoĭ. V rabote metodom molekuliarnoĭ dinamiki issledovali rol' soputstvuiushchikh mutatsiĭ v strukture b-laktamaz TEM-72, kotoraia soderzhit dve kliuchevye mutatsii (G238S, E240K) i dve soputstvuiushchie (Q39K, M182T). Pokazano, chto poiavlenie kliuchevykh mutatsiĭ ostatkov 238 i 240 privodilo k destabilizatsii belkovoĭ globuly, chto vyrazhalos' v uvelichenii podvizhnosti aminokislotnykh ostatkov. Soputstvuiushchie mutatsii ostatkov 182 i 39 vliiali na strukturu belka protivopolozhnym obrazom. Mutatsiia M182T uvelichivala ustoĭchivost' belkovoĭ globuly, a mutatsiia Q39K okazyvala destabiliziruiushchiĭ éffekt. Éta mutatsiia, po-vidimomu, sposobstvuet optimizatsii konformatsionnoĭ podvizhnosti b-laktamazy i mozhet vliiat' na rabotu fermenta.
Keywords: TEM-1; TEM-72; beta-lactamase; molecular dynamics; mutations; thermoresistance.