The role of Cys271 in conformational changes of arginine kinase |
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Authors: | Liu Na Wang Jin-Song Wang Wei-Dong Pan Ji-Cheng |
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Institution: | a College of Life Science, Hubei Normal University, Huangshi, Hubei 435002, PR China b Hubei Key Laboratory of Pollutant Analysis & Reuse Technology, PR China |
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Abstract: | Arginine kinase (AK), a crucial enzyme in energy metabolism, buffers cellular ATP levels by catalyzing the reversible phosphoryl transfer between ATP and arginine. To better understand the role of Cys271 in conformational changes of AK from greasyback shrimp (Metapenaeus ensis), we replaced the residue with serine and alanine. A detailed comparison of the catalytic activity and conformation was made between wild-type AK and the mutants by means of activity analysis, ultraviolet (UV) difference, fluorescence spectrum and size exclusion chromatography (SEC). The results indicated that the catalytic activity of the two mutants was gone. The substrates, arginine-ADP-Mg2+ could induce conformational changes, and additional NO3− could induce further changes in both the native enzyme and the variants. We speculated that Cys271 might be located in the hinge region between the two domains of AK and cause enzyme conformational changes upon addition of substrate. |
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Keywords: | WT-AK wild-type arginine kinase CK creatine kinase Cys271 cysteine of residue-271 SEC size exclusion chromatography TSAC transition state analog complex IPTG d-thiogalactoside" target="_blank">isopropyl-β-d-thiogalactoside ANS 1-anilinonaphtalene-8-sulfonate |
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