Shakil Shazi, Khan Asad Ullah
Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh-202002, Interdisciplinary Biotechnology Unit, Aligarh Muslim University Aligarh 202002, India.
Bioinformation. 2010 Apr 30;4(10):468-72. doi: 10.6026/97320630004468.
Extended-spectrum β-lactamases (ESBLs) are the bacterial enzymes that make them resistant to advanced-generation cephalosporins. CTXM enzymes (the most prevalent ESBL-type) target cefotaxime. Aims of the study were: Modelling of CTX-M enzyme from bla(CTX-M) sequences of clinical Escherichia coli isolatesDocking of cefotaxime with modelled CTX-M enzymes to identify amino acid residues crucial to their interaction To hypothesize a possible relationship between 'interaction energy of the docked enzyme-antibiotic complex' and 'minimum inhibitory concentration (MIC) of the antibiotic against the bacteria producing that enzyme'. Seven E. coli strains of clinical origin which were confirmed as PCR-positive for bla(CTX-M) were selected for the study. C600 cells harboring cloned bla(CTX-M) were tested for ESBL-production by double-disk-synergy test. BLAST analysis confirmed all the bla(CTX-M) genes as blaCTX-M-15. Four of the 7 strains were found to be clonally related. Modelling was performed using Swiss Model Server. Discovery Studio 2.0 (Accelrys) was used to prepare Ramachandran plots for the modelled structures. Ramachandran Z-scores for modelled CTX-M enzymes from E. coli strains D8, D183, D253, D281, D282, D295 and D296 were found to be -0.449, 0.096, 0.027, 0.043, 0.032, -1.249 and -1.107, respectively. Docking was performed using Hex 5.1 and the results were further confirmed by Autodock 4.0. The amino acid residues Asn 104, Asn132, Gly 227, Thr 235, Gly 236, and Ser237 were found to be responsible for positioning cefotaxime into the active site of the CTX-M-15 enzyme. It was found that cefotaxime MICs for the CTX-M-15-producers increased with the increasing negative interaction energy of the enzyme-antibiotic complex.
超广谱β-内酰胺酶（ESBLs）是使细菌对新一代头孢菌素产生耐药性的酶。CTX-M酶（最常见的ESBL类型）作用于头孢噻肟。本研究的目的是：根据临床大肠杆菌分离株的bla(CTX-M)序列对CTX-M酶进行建模；将头孢噻肟与建模的CTX-M酶对接，以确定对它们相互作用至关重要的氨基酸残基；推测“对接的酶-抗生素复合物的相互作用能”与“抗生素对产生该酶的细菌的最低抑菌浓度（MIC）”之间可能存在的关系。选择7株临床来源的大肠杆菌菌株，这些菌株经PCR检测bla(CTX-M)呈阳性用于本研究。通过双纸片协同试验检测携带克隆bla(CTX-M)的C600细胞的ESBL产生情况。BLAST分析证实所有bla(CTX-M)基因均为blaCTX-M-15。7株菌株中有4株被发现具有克隆相关性。使用瑞士模型服务器进行建模。使用Discovery Studio 2.0（Accelrys）为建模结构绘制拉氏图。发现来自大肠杆菌菌株D8、D183、D253、D281、D282、D295和D296的建模CTX-M酶的拉氏Z分数分别为-0.449、0.096、0.027、0.043、0.032、-1.249和-1.107。使用Hex 5.1进行对接，并通过Autodock 4.0进一步确认结果。发现氨基酸残基Asn 104、Asn132、Gly 227、Thr 235、Gly 236和Ser237负责将头孢噻肟定位到CTX-M-15酶的活性位点。发现对于产生CTX-M-15的菌株，头孢噻肟的MIC随着酶-抗生素复合物负相互作用能的增加而增加。
