An effective enzymatic assay for pH selectively measuring direct and total bilirubin concentration by using of CotA

Chengyu Zhang; Lin Zhu; Jiaxing Zhang; Wenhang Wang; Yan Zeng; Shengping You; Wei Qi; Rongxin Su; Zhimin He

doi:10.1016/j.bbrc.2021.01.094

An effective enzymatic assay for pH selectively measuring direct and total bilirubin concentration by using of CotA

Biochem Biophys Res Commun. 2021 Apr 2:547:192-197. doi: 10.1016/j.bbrc.2021.01.094. Epub 2021 Feb 19.

Authors

Chengyu Zhang¹, Lin Zhu¹, Jiaxing Zhang¹, Wenhang Wang², Yan Zeng³, Shengping You⁴, Wei Qi⁵, Rongxin Su⁶, Zhimin He⁷

Affiliations

¹ Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, PR China.
² Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin, 300072, PR China.
³ Tianjin Key Laboratory of Membrane Science and Desalination Technology, Tianjin University, Tianjin, 300072, PR China.
⁴ Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, PR China. Electronic address: ysp@tju.edu.cn.
⁵ Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, PR China; State Key Laboratory of Chemical Engineering, Tianjin University, Tianjin, 300072, PR China. Electronic address: qiwei@tju.edu.cn.
⁶ Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, PR China; State Key Laboratory of Chemical Engineering, Tianjin University, Tianjin, 300072, PR China.
⁷ Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin, 300072, PR China; State Key Laboratory of Chemical Engineering, Tianjin University, Tianjin, 300072, PR China. Electronic address: enzyme@tju.edu.cn.

PMID: 33618226
DOI: 10.1016/j.bbrc.2021.01.094

Abstract

In this study, we aimed to develop B. subtilis spore coat protein A (CotA) for the enzymatic determination of bilirubin. Firstly, molecular docking and oxidation kinetic analysis confirmed the feasibility of CotA for oxidizing bilirubin. Secondly, CotA showed pH-preferable oxidization performance to direct bilirubin (DB) in acidic conditions and an alkaline-catalytic oxidation capacity to total bilirubin (TB). Mechanism analysis results confirm that the conformational changes of CotA, DB and UB caused by pH changes are responsible for the selective oxidation of DB and TB by CotA. Then, CotA exhibits better structural characteristics and enzymatic performance than M. verrucaria-derived bilirubin oxidase (Mv-BOD). Besides, the strong anti-interference ability helps CotA adapt to complex catalytic environment in the detection of DB and TB. Our results prove that CotA can be used as a promising candidate bio-enzymatic detection reagent for DB and TB.

Keywords: Anti-interference; CotA; Direct bilirubin; Total bilirubin; pH-preferable oxidization.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Bacillus subtilis / enzymology*
Bacterial Proteins / chemistry
Bacterial Proteins / metabolism*
Bilirubin / analysis*
Bilirubin / metabolism
Enzyme Assays / methods*
Humans
Hydrogen-Ion Concentration
Laccase / chemistry
Laccase / metabolism*
Molecular Docking Simulation
Oxidation-Reduction

Substances

Bacterial Proteins
Laccase
Bilirubin