Chowdhury Trinath, Sarkar Manas, Chaudhuri Biswadeep, Chattopadhyay Brajadulal, Halder Umesh Chandra
Department of Physics, Jadavpur University, Kolkata, 700032, India.
J Biol Inorg Chem. 2015 Jul;20(5):791-803. doi: 10.1007/s00775-015-1266-2. Epub 2015 May 6.
A unique protein, bioremediase (UniProt Knowledgebase Accession No.: P86277), isolated from a hot spring bacterium BKH1 (GenBank Accession No.: FJ177512), has shown to exhibit silica leaching activity when incorporated to prepare bio-concrete material. Matrix-assisted laser desorption ionization mass spectrometry analysis suggests that bioremediase is 78% homologous to bovine carbonic anhydrase II though it does not exhibit carbonic anhydrase-like activity. Bioinformatics study is performed for understanding the various physical and chemical parameters of the protein which predicts the involvement of zinc encircled by three histidine residues (His94, His96 and His119) at the active site of the protein. Isothermal titration calorimetric-based thermodynamic study on diethyl pyrocarbonate-modified protein recognizes the presence of Zn(2+) in the enzyme moiety. Exothermic to endothermic transition as observed during titration of the protein with Zn(2+) discloses that there are at least two binding sites for zinc within the protein moiety. Addition of Zn(2+) regains the activity of EDTA chelated bioremediase confirming the presence of extra binding site of Zn(2+) in the protein moiety. Revival of folding pattern of completely unfolded urea-treated protein by Zn(2+) explains the participatory role of zinc in structural stability of the protein. Restoration of the λ max in intrinsic fluorescence emission study of the urea-treated protein by Zn(2+) similarly confirms the involvement of Zn in the refolding of the protein. The utility of bioremediase for silica nanoparticles preparation is observed by field emission scanning electron microscopy.
从温泉细菌BKH1（GenBank登录号：FJ177512）中分离出的一种独特蛋白质生物修复酶（UniProt知识库登录号：P86277），在用于制备生物混凝土材料时显示出硅浸出活性。基质辅助激光解吸电离质谱分析表明，生物修复酶与牛碳酸酐酶II有78%的同源性，尽管它不表现出碳酸酐酶样活性。为了解该蛋白质的各种物理和化学参数进行了生物信息学研究，该研究预测在蛋白质活性位点上有三个组氨酸残基（His94、His96和His119）环绕的锌的参与。基于等温滴定量热法对焦碳酸二乙酯修饰的蛋白质进行的热力学研究识别出酶部分中存在Zn(2+)。在用Zn(2+)滴定蛋白质过程中观察到的放热到吸热转变表明，蛋白质部分内至少有两个锌结合位点。添加Zn(2+)可恢复经EDTA螯合的生物修复酶的活性，证实蛋白质部分中存在额外的Zn(2+)结合位点。Zn(2+)使完全展开的尿素处理蛋白质的折叠模式恢复，这解释了锌在蛋白质结构稳定性中的参与作用。Zn(2+)在尿素处理蛋白质的内在荧光发射研究中恢复λ max同样证实了Zn参与蛋白质的重折叠。通过场发射扫描电子显微镜观察到生物修复酶在制备二氧化硅纳米颗粒方面的效用。
