Structural and catalytic alteration of sarcosine oxidase through reconstruction with coenzyme-like ligands

Publication date: Available online 13 January 2017 Source:Journal of Molecular Catalysis B: Enzymatic Author(s): Yu Xin, Mengling Zheng, Qing Wang, Liushen Lu, Ling Zhang, Yanjun Tong, Wu Wang A sarcosine oxidase (SOX) gene from Bacillus sp. (AY626822.2) was expressed in Escherichia coli BL21 (DE3) in the form of inclusion bodies. A 3D model of SOX was then built and refined, and molecular docking was used to investigate the interactions between SOX and natural or coenzyme-like ligands, including flavin adenine dinucleotide (FAD); flavin mononucleotide (FMN); riboflavin; isoalloxazine; 7-methyl-8-chloro-10-(1′-D-ribityl) isoalloxazine (7-M-8-C); 7-bromo-8-methyl-10-(1′-D-ribityl) isoalloxazine (7-B-8-M); 7-methyl-8-bromo-10-(1′-D-ribityl) isoalloxazine (7-M-8-B); 7-chloro-8-ethyl-10-(1′-D-ribityl) isoalloxazine (7-C-8-E); 7,8-diethyl-10-(1′-D-ribityl) isoalloxazine (7,8-D); and 3-methyl-7,8-dimethyl-10-(1′-D-ribityl) isoalloxazine (3-M-7,8-D). Unfolded SOX was extracted from inclusion bodies, and reconstructed with these ligands via a refolding process. The reconstructed enzymes were then subjected to structural and catalytic analysis. After structural simulation, refinement, and molecular docking, all ligands were able to recognize the coenzyme site of SOX. In addition, when the position 7- or 8-site of the compounds was modified, new pi-cation/sigma interactions were formed in the SOX-ligand complex. Fluorescent detection revealed that all the ligands c...
Source: Journal of Molecular Catalysis B: Enzymatic - Category: Biochemistry Source Type: research