A new target region for changing the substrate specificity of amine transaminases

• Published in: Scientific reports Vol. 5; no. 1; p. 10753
• Main Authors: Guan, Li-Jun, Ohtsuka, Jun, Okai, Masahiko, Miyakawa, Takuya, Mase, Tomoko, Zhi, Yuehua, Hou, Feng, Ito, Noriyuki, Iwasaki, Akira, Yasohara, Yoshihiko, Tanokura, Masaru
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01.06.2015; Nature Publishing Group
• Subjects: 631/1647/338/469; 631/45/603; 631/45/607/1172; 82/16; 82/80; 82/83; Amines - metabolism; Amino Acid Substitution; Binding Sites; Biocatalysts; Catalytic Domain; Conformation; Conserved Sequence; Crystal structure; Diabetes mellitus; Humanities and Social Sciences; Models, Molecular; multidisciplinary; Mutation; Protein Binding; Protein Conformation; Protein Multimerization; Science; Structure-Activity Relationship; Substrate Specificity; Surface properties; Transaminases - chemistry; Transaminases - genetics; Transaminases - metabolism
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/srep10753

( R )-stereospecific amine transaminases ( R -ATAs) are important biocatalysts for the production of ( R )-amine compounds in a strict stereospecific manner. An improved R -ATA, ATA-117-Rd11, was successfully engineered for the manufacture of sitagliptin, a widely used therapeutic agent for type-2 diabetes. The effects of the individual mutations, however, have not yet been demonstrated due to the lack of experimentally determined structural information. Here we describe three crystal structures of the first isolated R -ATA, its G136F mutant and engineered ATA-117-Rd11, which indicated that the mutation introduced into the 136 th residue altered the conformation of a loop next to the active site, resulting in a substrate-binding site with drastically modified volume, shape and surface properties, to accommodate the large pro-sitagliptin ketone. Our findings provide a detailed explanation of the previously reported molecular engineering of ATA-117-Rd11 and propose that the loop near the active site is a new target for the rational design to change the substrate specificity of ATAs.