The Crystal Structure and Molecular Docking of CYP76AH3 in the Tanshinone Biosynthesis Pathway / 中国生物化学与分子生物学报

ABSTRACT

Salvia miltiorrhiza is widely used in the treatment of the angina pectoris, coronary heart disease and myocardial infarction. CYP76AH3 is the key P450 enzyme, locating at the branch point of the tanshinone synthesis pathway. The crystal structural study and key amino acid analysis are of great significance for synthetic biology study on tanshinone. But it is always a challenge for scientists to carry out protein purification, crystallization and crystal structural studies on transmembrane type Ⅱ P450 enzymes. In this study, the prokaryotic expression plasmid was generated, and the high-purity target protein was purified. CYP76AH3 was successfully crystallized, and the crystal structure was solved.After docking range was determined by Cavityplus analysis, molecular docking with Discovery Studio was conducted. The docking result indicated that Gly298 and Asp294 had hydrogen bond interaction with the substrate, while Phe479, Leu367 and Leu293 had hydrophobic interaction with the substrate.In addition, the effect of mutations at the key amino acids on the protein structure stability was predicted throung point mutation simulation. This study would provide a target for protein engineering of CYP76AH3 and lay a foundation for the study of synthesis biology on tanshinones.