The bitter peptide fraction present in casein hydrolysates obtained by using three proteases (subtilisin, papain and trypsin) was treated with aminopeptidase T from 7'hennus aquaticus YT-1. The bitterness of the bitter peptide fraction could be decreased, and it sometimes disappeared completely, with an increase in free amino acids. The percentages of total free amino acids released from each bitter peptide fraction (subtilisin, papain and trypsin) by aminopeptidase digestion for 20 hr were approximately ll%, 8.7%, and 6.5%, respectively. Bitter peptide (a,l-CN f91-100) was isolated from a tryptic hydrolysate of casein by HPLC, its threshold value of bitterness being 2.9 ppm (w/v). The peptide (%l-CN f96-100) obtained from the amino peptidase digestion of this bitter peptide showed no bitterness.

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A thermostable aminopeptidase, called aminopeptidase T, from the extract of Thermus aquaticus YT-1 was purified and characterized. The enzyme had a dimeric structure, its relative molecular mass being 108,000 by gel filtration, and 48,000 by SDS-PAGE. The optimum pH of the enzymeactivity was in the range of 8.5 to 9.0. The enzymewas significantly thermostable as it still retained 60%of its original activity even after heat treatment for 20 hr at 80°C. The enzyme activity was inhibited by metal-chelating agents. The enzyme had a low substrate specificity.

Carboxypeptidase (CPase) Taq possesses the His-Glu-X-X-His sequence, which is the consensus sequence in the active site of zinc-dependent endopeptidases and amino-peptidases, at positions 276-280. Amino acid replacement of the conserved His and Glu drastically diminished the activity of CPase Taq, and the zinc content of the enzyme was also greatly reduced when either of the two His residues was replaced with Arg or Tyr. The results indicate that this sequence actually functions as the active site in CPase Taq, showing that CPase Taq is a novel type of zinc-dependent CPase that possesses the His-Glu-X-X-His active-site motif.