title: Characterization of (R)-selective amine transaminases identified by in silico motif sequence blast.
abstract: Compared to (S)-selective amine transaminase ((S)-AT), the (R)-selective counterpart ((R)-AT) has been less studied. As such, a simplified "Motif Sequence Blast" search (Höhne et al. Nat Chem Biol 6:807-813, 2010) was carried out to identify new (R)-ATs from the protein databases. The combined conserved sequence motifs of (R)-ATs based on the previous in silico method of predicting (R)-selective amine transaminase were used as the template sequence for BLASTP search at default settings in NCBI, and six candidate sequences were identified. These putative (R)-AT genes were synthesized and overexpressed in Escherichia coli. Among them, five new (R)-ATs were expressed as soluble protein and showed unusual substrate specificity and high stereoselectivity. Furthermore, several unnatural amino acids, such as D-alanine, D-2-aminobutyric acid, and D-norvaline, were synthesized via the (R)-AT-catalyzed amino transfer reaction to the corresponding keto acids. Optically pure (S)-amines were also obtained by kinetic resolution of racemic amines catalyzed with these new (R)-ATs. Therefore, the Motif Sequence Blast search offers a quick and effective method for in silico identification of new (R)-ATs, and the newly identified (R)-ATs are attractive additions to the toolbox of (R)-ATs for further study and industrial application.
authors.affiliation: National Engineering Laboratory for Industrial Enzymes and Tianjin Engineering Center for Biocatalytic Technology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China.
authors.forename: Jinju
authors.initials: J
authors.lastname: Jiang
authors.forename: Xi
authors.initials: X
authors.lastname: Chen
authors.forename: Dalong
authors.initials: D
authors.lastname: Zhang
authors.forename: Qiaqing
authors.initials: Q
authors.lastname: Wu
authors.forename: Dunming
authors.initials: D
authors.lastname: Zhu
mesh_terms: D000409:Alanine / Q000378:metabolism; D000613:Aminobutyrates / Q000378:metabolism; D032661:Cordyceps / Q000235:genetics / Q000378:metabolism; D030562:Databases, Protein; D005670:Fusarium / Q000235:genetics / Q000378:metabolism; D007651:Keto Acids / Q000378:metabolism; D055330:Nectria / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D000637:Transaminases / Q000235:genetics* / Q000378:metabolism*; D014242:Trichoderma / Q000235:genetics / Q000378:metabolism; D014633:Valine / Q000031:analogs & derivatives / Q000378:metabolism
chemical_list: D000613:Aminobutyrates; D007651:Keto Acids; C012223:alpha-aminobutyric acid; C005313:norvaline; D000637:Transaminases; D014633:Valine; D000409:Alanine