Method of using N-acetyl-2,3-Didehydroleucine acylase for the preparation of D- or L-tryptophyl glycine, D- or L-tryptophyl-D-methionine or L-tryptophyl-D-cysteine

A method for the enzymatic conversion of D- or L-tryptophane amide and glycine to the dipeptide D- or L-tryptophyl glycine, of D- or L-tryptophane amide and D-methionine to D- or L-tryptophy-D-methionine or of L-tryptophane amine and D-cysteine to the dipeptide L-tryptophyl-D-cysteine, said method comprising carrying out the conversion in the presence of a N-acetyl-2,3-didehydroleucine acylase, having the following characteristics

• 1) reactivity:
• it splits off the acetyl group from N-acetyl-2,3-didehydroleucine, at which time acetic acid and, after consecutive chemical reactions, 2-keto-4-methyl valeric acid and ammonia arise as end products;
• 2) Substrate specificity;
• it hydrolyses both various N-acetyl-2,3-didehydroamino acids such as N-acetyl-2,3-didehydrovaline, N-acetyl-2,3-didehydroisoleucine, 2-acetylamino cinnamic acid and 2-acetylamino acrylic acid as well as amino acid amides such as D- and L-tryptophane amide, D- and L-leucine amide and L-methionine amide but not 2,3-saturated N-acetylamino carboxylic acids such as N-acetyl leucine or N-acetyl valine;
• 3) optimum pH:
• the optimum pH is 9.3+1;
• 4) pH stability:
• it exhibits good stability in a pH range between 9 and 10;
• 5) optimum temperature:
• the optimum temperature is 55.degree. C. at a pH of 9;
• 6) temperature resistance:
• at 50.degree. C., 90% residual activity can still be demonstrated after 30 minutes incubation;
• 7) influences of inhibitors and activators:
• Inhibitors of serine proteases, especially phenylmethane sulfonylfluoride (0.001 mM), exert an inhibitory action, glycine accelerates the substrate splitting as a function of the concentration;
• 8) molecular weight:
• the molecular weight is approximately 60000;
• 9) subunits:
• the molecule consists of only one unit;
• 10) K-value:
• the K-value for the substrate N-acetyl-2,3-didehydroleucine is 4.5 mM (30.degree. C., 0.1M glycine buffer, pH 9).