Dalal Vikram, Dhankhar Poonam, Singh Vishakha, Singh Vishakha, Rakhaminov Gaddy, Golemi-Kotra Dasantila, Kumar Pravindra
Department of Biotechnology, IIT Roorkee, Roorkee, Uttrakhand, 247667, India.
Department of Biology, York University, 4700 Keele Street, Toronto, Canada.
Protein J. 2021 Apr;40(2):148-165. doi: 10.1007/s10930-020-09953-6. Epub 2021 Jan 9.
Staphylococcus aureus is resistant to β-lactam antibiotics and causes several skin diseases to life-threatening diseases. FmtA is found to be one of the main factors involved in methicillin resistance in S. aureus. FmtA exhibits an esterase activity that removes the D-Ala from teichoic acid. Teichoic acids played a significant role in cell wall synthesis, cell division, colonization, biofilm formation, virulence, antibiotic resistance, and pathogenesis. The virtual screening of drug molecules against the crystal structure of FmtA was performed and the binding affinities of top three molecules (ofloxacin, roflumilast, and furazolidone) were predicted using molecular docking. The presence of positive potential and electron affinity regions in screened drug molecules by DFT analysis illustrated that these molecules are reactive in nature. The protein-ligand complexes were subjected to molecular dynamics simulation. Molecular dynamics analysis such as RMSD, RMSF, Rg, SASA, PCA, and FEL results suggested that FmtA-drug(s) complexes are stable. MM-GBSA binding affinity and QM/MM results (ΔG, ΔH, and ΔS) revealed that active site residues (Ser127, Lys130, Tyr211, Asp213, and Asn343) of FmtA played an essential for the binding of the drug(s) to form a lower energy stable protein-ligand complexes. FmtA was purified using cation exchange and gel filtration chromatography. Fluorescence spectroscopy and circular dichroism results showed that interactions of drugs with FmtA affect the tertiary structure and increase the thermostability of the protein. The screened molecules need to be tested and could be further modified to develop the antimicrobial compounds against S. aureus.
金黄色葡萄球菌对β-内酰胺抗生素具有抗性，并会引发多种从皮肤病到危及生命的疾病。FmtA被发现是金黄色葡萄球菌耐甲氧西林的主要因素之一。FmtA具有酯酶活性，可从磷壁酸中去除D-丙氨酸。磷壁酸在细胞壁合成、细胞分裂、定植、生物膜形成、毒力、抗生素抗性和发病机制中发挥着重要作用。针对FmtA的晶体结构进行了药物分子的虚拟筛选，并使用分子对接预测了前三个分子（氧氟沙星、罗氟司特和呋喃唑酮）的结合亲和力。通过密度泛函理论（DFT）分析筛选出的药物分子中存在正电势和电子亲和力区域，表明这些分子在本质上具有反应性。对蛋白质-配体复合物进行了分子动力学模拟。诸如均方根偏差（RMSD）、均方根波动（RMSF）、回旋半径（Rg）、溶剂可及表面积（SASA）、主成分分析（PCA）和自由能景观（FEL）等分子动力学分析结果表明，FmtA-药物复合物是稳定的。分子力学广义玻恩表面面积（MM-GBSA）结合亲和力和量子力学/分子力学（QM/MM）结果（ΔG、ΔH和ΔS）表明，FmtA的活性位点残基（Ser127、Lys130、Tyr211、Asp213和Asn343）对于药物结合形成能量更低的稳定蛋白质-配体复合物至关重要。使用阳离子交换和凝胶过滤色谱法纯化了FmtA。荧光光谱和圆二色性结果表明，药物与FmtA的相互作用会影响三级结构并提高蛋白质的热稳定性。筛选出的分子需要进行测试，并可进一步修饰以开发针对金黄色葡萄球菌的抗菌化合物。
