D-aminoacylase and gene encoding the same

#The present invention relates to a D-aminoacylase, a gene encoding the D-aminoacylase, and a method for producing D-amino acids using the D-aminoacylase.

#Enzymes have excellent catalytic functions with substrate specificity, reaction specificity, and stereospecificity. Stereospecificity of enzymes, with some exceptions, are nearly absolute.

#Recent precise research has increased the importance of optically active substances for use in drugs, pesticides, feeds, and perfumes. Since optical isomers sometimes have quite different biological activities, techniques for specifically obtaining the isomer are important. For example, D(R)-form thalidomide has no teratogenic activity, but its L(S)-form shows strong teratogenicity. The practical use of thalidomide racemate caused the drug injury incidents by thalidomide. Furthermore, if one enantiomer shows an effective biological activity, the other enantiomer may sometimes have no activity, rather, it may competitively inhibit the activity of the effective enantiomer because of the coexistence of both enantiomers. As a result, the biological activity of the racemate is reduced to half or less of the activity of the effective enantiomer. Accordingly, it is industrially important to obtain (synthesize or optically resolve) optically pure enantiomers.

#The present invention relates to an isolated polynucleotide encoding a D-aminoacylase and homologues thereof.

#An “isolated polynucleotide” is a polynucleotide the structure of which is not identical to that of any naturally occurring polynucleotide or to that of any fragment of a naturally occurring genomic polynucleotide spanning more than three separate genes. The term therefore includes, for example, (a) a DNA which has the sequence of part of a naturally occurring genomic DNA molecule in the genome of the organism in which it naturally occurs; (b) a polynucleotide incorporated into a vector or into the genomic DNA of a prokaryote or eukaryote in a manner such that the resulting molecule is not identical to any naturally occurring vector or genomic DNA; (c) a separate molecule such as a cDNA, a genomic fragment, a fragment produced by polymerase chain reaction (PCR), or a restriction fragment; and (d) a recombinant nucleotide sequence that is part of a hybrid gene, i.e., a gene encoding a fusion protein. Specifically excluded from this definition are polynucleotides of DNA molecules present in mixtures of different (i) DNA molecules, (ii) transfected cells, or (iii) cell clones; e.g., as these occur in a DNA library such as a cDNA or genomic DNA library.

#The polynucleotide encoding the D-aminoacylase of the present invention comprises, for example, the nucleotide sequence of SEQ ID NO: 1. The nucleotide sequence of SEQ ID NO: 1 encodes a polypeptide comprising the amino acid sequence of SEQ ID NO: 2. The polypeptide comprising this amino acid sequence constitutes preferred embodiments of the D-aminoacylase of the present invention. An amino acid is not always encoded by a single codon; because of degeneracy, there can be multiple codons corresponding to an amino acid, which is well known to those skilled in the art. Therefore, nucleotide sequences encoding the amino acid sequence shown in SEQ ID NO: 2 comprise not only the nucleotide sequence of SEQ ID NO: 1 but also any nucleotide sequence comprising other different codons as long as it encodes the amino acid sequence of SEQ ID NO: 2.