A comparison of the partial amino-acid sequence of neutral protease A from Bacillus subtilis with the structure of thermolysin (EC 3.4.24.4) from Bacillus thermoproteolyticus reveals that these two proteins are homologous. Of 171 residues placed in neutral protease (54% of the sequence), 83 residues (49%) occur in identical positions in thermolysin, and include nine of the 13 residues previously identified as components of the active site of thermolysin. This similarity provides support for the hypothesis that the two enzymes have similar three-dimensional structures and a common mechanism of action. Since these enzymes differ markedly in their resistance to heat inactivation, a comparison of their structures may eventually provide a chemical basis for explaining the differences in their thermal stability. The metalloendopeptidases comprise a group of proteolytic enzymes that share a number of functional properties; these include a pH optimum near 7.0, inhibition by chelating agents-and resistance to inactivation by diisopropylphosphofluoridate or thiol reagents (1). Such "neutral proteases" have been isolated from a wide variety of sources that include bacteria, fungi, and more recently the brush border cells of rabbit kidney (1, 2). Apart from the notable exception of the amino-acid sequence (3) and three-dimensional structure (4) of thermolysin (EC 3.4.24.4), relatively little is known concerning the structural features of the metalloendopeptidases and their relation to function.Various studies have indicated that a neutral protease isolated from the culture medium of Bacilllus subtilis NRRLB3411 appears to be quite similar to thermolysin in amino-acid composition (5), kinetic behavior (6), and susceptibility to inhibitors (1, 7), which suggests that these two enzymes might be homologous. This investigation was undertaken to develop sequence data that would clarify the possible structural interrelationships between the neutral metalloendopeptidases, and in particular demonstrate that the neutral protease A from B. subtilis and the thermostable protease, thermolysin, are homologous proteins.

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