Catalytic performance and molecular dynamic simulation of immobilized CC bond hydrolase based on carbon nanotube matrix

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• 作者：Hao Zhou ; Yuanyuan Qu ; ^{qyy@dlut.edu.cn" class="auth_mail} ; Chunlei Kong ; Duanxing Li ; E. Shen ; Qiao Ma ; Xuwang Zhang ; Jingwei Wang ; Jiti Zhou
• 关键词：AFM ; atomic force microscopy ; CD ; circular dichroism ; CNT ; carbon nanotube ; DMF ; dimethylformamide ; EDC ; N-ethyl-N&prime ; -(3-(dimethyllamino) propyl) carbodiimide hydrochloride ; MD ; molecular dynamics ; MES ; 2-(N-morpholino) ethanesulfonic acid ; MWCNT ; multi wall carbon nanotube ; NHS ; N-hydroxysuccinimide ; PNPA ; p-nitrophenyl acetate ; PNPBen ; p-nitrophenyl benzoate ; SASA ; solvent accessible surface area ; SDS-PAGE ; sodium dodecyl sulfate polyacrylamide gel electrophoresis ; SWCNT ; single wall carbo
• 刊名：Colloids and Surfaces B: Biointerfaces
• 出版年：1 April, 2014
• 年：2014
• 卷：116
• 期：Complete
• 页码：365-371
• 全文大小：1896 K

文摘

Carbon nanotube (CNT) has been proved to be a kind of novel support for enzyme immobilization. In this study, we tried to find the relationship between conformation and catalytic performance of immobilized enzyme. Two Ccript>cript>C bond hydrolases BphD and MfphA were immobilized on CNTs (SWCNT and MWCNT) via physical adsorption and covalent attachment. Among the conjugates, the immobilized BphD on chemically functionalized SWCNT (BphD-CSWCNT) retained the highest catalytic efficiency (k_cat/K_m value) compared to free BphD (92.9%). On the other hand, when MfphA bound to pristine SWCNT (MfphA-SWCNT), it was completely inactive. Time-resolved fluorescence spectrum indicated the formation of static ground complexes during the immobilization processes. Circular dichroism (CD) showed that the secondary structures of immobilized enzymes changed in varying degrees. In order to investigate the inhibition mechanism of MfphA by SWCNT, molecular dynamics simulation was employed to analyze the adsorption process, binding sites and time evolution of substrate tunnels. The results showed that the preferred binding sites (Trp201 and Met81) of MfphA for SWCNT blocked the main substrate access tunnel, thus making the enzyme inactive. The 鈥渢unnel-block鈥?should be a novel possible inhibition mechanism for enzyme-nanotube conjugate.