The present disclosure relates to polypeptides having transaminase activity, polynucleotides encoding the polypeptides, and methods of using the polypeptides.

The present disclosure provides engineered transaminase enzymes having improved properties as compared to a naturally occurring wild-type transaminase enzyme. Also provided are polynucleotides encoding the engineered transaminase enzymes, host cells capable of expressing the engineered transaminase enzymes, and methods of using the engineered transaminase enzymes to synthesize a variety of chiral compounds.

The present invention relates to the production of optically pure secondary amines, which can be used as intermediate products in a synthesis of for instance pharmaceutical products.

The present disclosure relates to polypeptides having transaminase activity, polynucleotides encoding the polypeptides, and methods of using the polypeptides.

The present disclosure relates to polypeptides having transaminase activity, polynucleotides encoding the polypeptides, and methods of using the polypeptides.

A Pseudomonas putida -aminotransferase gene with broad substrate spectrum and an application thereof are provided. According to the disclosure, codon optimization is carried out on an -transaminase gene, and the nucleotide sequence of the -transaminase gene is shown in SEQ ID NO: 1. The crude enzyme solution, crude enzyme powder and pure enzyme of -aminotransferase prepared from -aminotransferase gene can be used as an enzyme preparation to catalyze the transamination reaction of substrates to prepare chiral amino alcohols and chiral amines. The substrate spectrum of the recombinant -transaminase obtained by the disclosure is broad, and 15 kinds of chiral amino alcohols and chiral amines with high added value are asymmetrically catalyzed and synthesized; in particular, 4-hydroxy-2-butanone is catalyzed to produce pharmaceutical intermediate (R)-3-amino-1-butanol.

The present disclosure provides engineered transaminase polypeptides for the production of amines, polynucleotides encoding the engineered transaminases, host cells capable of expressing the engineered transaminases, and methods of using the engineered transaminases to prepare compounds useful in the production of active pharmaceutical agents.

The present invention discloses a transaminase mutant and application thereof in preparation of sitagliptin intermediates, the transaminase mutant is obtained by substitution of tyrosine with proline at position 74, substitution of glutamic acid with aspartic acid at position 228, substitution of leucine with alanine at position 254 and substitution of methionine with threonine at position 290 of the amino acid sequence shown in SEQ ID NO: 2. The present invention uses wet cells or a purified transaminase as a biocatalyst and a sitagliptin precursor ketone or a prochiral carbonyl compound as a substrate to prepare a sitagliptin intermediate or a sitagliptin ester intermediate; the total yield of the method reaches about 82%, and e.e. value of the product reaches 99%.

The present invention discloses an -transaminase mutant based on ancestral sequence reconstruction and relates to the technical field of molecular biology. The amino acid sequence of said -transaminase mutant derived from mutation of -transaminase from Aspergillus terreus, is as shown in SEQ ID NO. 4 or SEQ ID NO. 6. Compared with the wild-type enzyme, the half-lives of the -transaminase mutants are all above 24 h, while the half-life of the wild-type is only 6.90 min. The half-inactivation temperature of the mutants are 49.00 C. and 49.03 C., respectively, which are about 11 C. higher than that of the wild-type (37.89 C.), such that the thermal stability is significantly improved.

Provided is an omega-transaminase mutant acquired by a single-point mutation or multi-point mutation at positions 275, 115, and 97 of the amino acid sequence set forth in SEQ ID NO. 2. The transaminase mutant is derived from Aspergillus lentulus. It catalyzes bioreactions with a ketone precursor of a sitagliptin intermediate as the substrate, isopropylamine as the amino donor, pyridoxal phosphate as the coenzyme, and a protonic polar solvent as the cosolvent, thus separating and purifying sitagliptin or the sitagliptin intermediate with high optical purity.