Crystal structure of CotA laccase complexed with 2, 2‐azinobis‐(3‐ethylbenzothiazoline‐6‐sulfonate) at a novel binding site. Issue 4 (1st April 2016)

• Record Type: Journal Article
• Authors: Liu, Zhongchuan; Xie, Tian; Zhong, Qiuping; Wang, Ganggang
• Is Part Of: Acta crystallographica. Volume 72:Issue 4(2016:Apr.)
• Page Start: 328
• Page End: 335
• Publication Date: 2016-04-01
• Subjects: 2, 2‐azinobis‐(3‐ethylbenzothiazoline‐6‐sulfonate) -- laccase -- mutagenesis -- substrate binding -- protein engineering; Crystallography -- Periodicals; Crystals -- Periodicals; 548
• DOI: 10.1107/S2053230X1600426X
• Languages: English
• ISSNs: 2053-230X

Abstract

Abstract : The crystal structure of CotA complexed with 2, 2‐azinobis‐(3‐ethylbenzothiazoline‐6‐sulfonate) in a hole motif has been solved; this novel binding site could be a potential structure‐based target for protein engineering of CotA laccase. Abstract : The CotA laccase from Bacillus subtilis is an abundant component of the spore outer coat and has been characterized as a typical laccase. The crystal structure of CotA complexed with 2, 2‐azinobis‐(3‐ethylbenzothiazoline‐6‐sulfonate) (ABTS) in a hole motif has been solved. The novel binding site was about 26 Å away from the T1 binding pocket. Comparison with known structures of other laccases revealed that the hole is a specific feature of CotA. The key residues Arg476 and Ser360 were directly bound to ABTS. Site‐directed mutagenesis studies revealed that the residues Arg146, Arg429 and Arg476, which are located at the bottom of the novel binding site, are essential for the oxidation of ABTS and syringaldazine. Specially, a Thr480Phe variant was identified to be almost 3.5 times more specific for ABTS than for syringaldazine compared with the wild type. These results suggest this novel binding site for ABTS could be a potential target for protein engineering of CotA laccases.