Polley Soumitra, Seal Soham, Mahapa Avisek, Jana Biswanath, Biswas Anindya, Mandal Sukhendu, Sinha Debabrata, Sau Keya, Sau Subrata
Department of Biochemistry, Bose Institute, Kolkata, West Bengal, India.
Department of Biotechnology, Haldia Institute of Technology, Haldia, West Bengal, India.
Bioinformation. 2017 Mar 31;13(3):78-85. doi: 10.6026/97320630013078. eCollection 2017.
Cyclophilins, a class of peptidyl-prolyl cis-trans isomerase (PPIase) enzymes, are inhibited by cyclosporin A (CsA), an immunosuppressive drug. Staphylococcus aureus Newman, a pathogenic bacterium, carries a gene for encoding a putative cyclophilin (SaCyp). SaCyp shows significant homology with other cyclophilins at the sequence level. A three-dimensional model structure of SaCyp harbors a binding site for CsA. To verify whether SaCyp possesses both the PPIase activity and the CsA binding ability, we have purified and investigated a recombinant SaCyp (rCyp) using various in vitro tools. Our RNase T1 refolding assay indicates that rCyp has a substantial extent of PPIase activity. rCyp that exists as a monomer in the aqueous solution is truly a cyclophilin as its catalytic activity specifically shows sensitivity to CsA. rCyp appears to bind CsA with a reasonably high affinity. Additional investigations reveal that binding of CsA to rCyp alters its structure and shape to some extent. Both rCyp and rCyp-CsA are unfolded via the formation of at least one intermediate in the presence of guanidine hydrochloride. Unfolding study also indicates that there is substantial extent of thermodynamic stabilization of rCyp in the presence of CsA as well. The data suggest that rCyp may be exploited to screen the new antimicrobial agents in the future.
亲环蛋白是一类肽基脯氨酰顺反异构酶（PPIase），可被免疫抑制药物环孢素A（CsA）抑制。金黄色葡萄球菌纽曼菌株是一种病原菌，携带一个编码假定亲环蛋白（SaCyp）的基因。SaCyp在序列水平上与其他亲环蛋白具有显著同源性。SaCyp的三维模型结构含有一个CsA结合位点。为了验证SaCyp是否同时具有PPIase活性和CsA结合能力，我们使用各种体外工具对重组SaCyp（rCyp）进行了纯化和研究。我们的核糖核酸酶T1重折叠试验表明rCyp具有相当程度的PPIase活性。在水溶液中以单体形式存在的rCyp确实是一种亲环蛋白，因为其催化活性对CsA具有特异性敏感性。rCyp似乎以相当高的亲和力结合CsA。进一步的研究表明，CsA与rCyp的结合在一定程度上改变了其结构和形状。在盐酸胍存在下，rCyp和rCyp-CsA都通过形成至少一种中间体而展开。展开研究还表明，在CsA存在下，rCyp也有相当程度的热力学稳定性。数据表明，rCyp未来可能被用于筛选新型抗菌剂。
