Siemann Stefan, Brewer Dyanne, Clarke Anthony J, Dmitrienko Gary I, Lajoie Gilles, Viswanatha Thammaiah
Department of Chemistry, University of Waterloo, 200 University Ave. W, Waterloo, ON, Canada N2L 3G1.
Biochim Biophys Acta. 2002 Jul 3;1571(3):190-200. doi: 10.1016/s0304-4165(02)00258-1.
Metallo-beta-lactamases have attracted considerable attention due to their role in microbial resistance to beta-lactam antibiotics. IMP-1, the binuclear Zn-dependent beta-lactamase produced by Pseudomonas aeruginosa and other microorganisms, is of particular interest in view of its increasing prevalence. An examination of the susceptibility of IMP-1 to inactivation by six different divalent metal ion chelators has revealed that all except Zincon cause inhibition by forming a complex with the holoenzyme. Exposure of the enzyme to dipicolinic acid (DPA), the most potent inhibitor, results in the production of the mononuclear Zn form of the protein as determined by electrospray ionization mass spectrometry (ESI-MS) under nondenaturing conditions. This mononuclear Zn species was found to be catalytically competent. Studies with the chromophoric chelator 4-(2-pyridylazo)resorcinol (PAR) show that the two zinc centers in IMP-1 differ in their accessibility, a feature that could be overcome in the presence of guanidine hydrochloride (GdnHCl, 1.5 M).
金属β-内酰胺酶因其在微生物对β-内酰胺抗生素耐药性中的作用而备受关注。IMP-1是由铜绿假单胞菌和其他微生物产生的双核锌依赖性β-内酰胺酶，鉴于其日益增加的流行率，尤其受到关注。对IMP-1被六种不同二价金属离子螯合剂失活的敏感性研究表明，除锌试剂外，其他螯合剂均通过与全酶形成复合物而导致抑制作用。将该酶暴露于最有效的抑制剂吡啶二甲酸（DPA）下，通过非变性条件下的电喷雾电离质谱（ESI-MS）测定，结果产生了蛋白质的单核锌形式。发现这种单核锌物种具有催化活性。用发色螯合剂4-（2-吡啶偶氮）间苯二酚（PAR）进行的研究表明，IMP-1中的两个锌中心在可及性方面存在差异，在存在盐酸胍（GdnHCl，1.5 M）的情况下可以克服这一特征。
