Recombinant decarbamylases for producing d-.alpha.-amino acids

Abstract

Decarbamylases are provided capable of producing D-.alpha.-amino acids by hydrolysis of N-carbamyl-D-.alpha.-amino acids. A source of the decarbamylases is recombinant microorganisms produced by gene manipulation methods. Decarbamylases having improved thermostability can be obtained in which amino acids at a thermostability-related site of a natural decarbamylase have been replaced with other amino acids by mutating a DNA fragment encoding the natural decarbamylase. Recombinant DNA is obtained from a vector DNA and a DNA fragment encoding a natural decarbamylase where the nucleic acid sequence encoding an amino acid at a thermostability-related site is replaced with a nucleic acid sequence encoding another amino acid. The recombinant DNA is used to produce transformants that produce thermostable decarbamylases. The decarbamylases may be immobilized in purified, partially-purified or crude form or in the form of decarbamylase-containing microbial cells on a support such as a polymer having ion exchange groups or covalent bonding groups. The presence of an antioxidant such as dithiothreitol results in an immobilized decarbamylase preparation that can be repeatedly used 30 times or more.