Carboxypeptidase inhibitors preparation

Gray and coworkers " have prepared copper(II) carboxypeptidase A. Cu"CPA, and compared its spectrum, that of the enzyme with inhibitor present, and those of several other copperdl) complexes with nitrogen and oxygen ligating atoms. Some of these data together with the geometry about the copper ion are ... 

It is sometimes extremely difficult to repeat a biochemical experiment, and the preparation of the crystalline form which we studied ( a= 51.41 A, c = 47.19 A and p = 97°35 ) was just such a problem. Only about two of some thirty preparations crystallized with these cell dimensions. On the other hand most preparations, including the commercial preparation, give cell dimensions of a= 50.9 A, b = 57.9 A, c = 45.0 A and p = 94°40. The activities of these two forms are very different in the solid state, 1/3 and 1/300 of that of the material in solution Proc. Nat. Acad. Sci. 70, 3797 (1973)). The arsanilzao Tyr 248 derivative of the former crystals behaves like the material in solution, but this derivative of the latter crystals is quite different from the material in solution. Vallee continues to discoimt the results of our crystallography, done on the former unit cell, on the basis of his results on the commercial form, which most likely has the latter unit cell. Probably the main difference is in the intermolecular contacts in the two crystalline forms. Since the crystals of the X-ray study are active, the conclusion of this work should be tested on that basis. Our present plan (1977) is to elucidate the structures of the protein inhibitor of carboxypeptidase A isolated from potatoes, and of its complex with the enzyme in the hope that atomic displacements toward the transition state can be recognized. 

In the last two years inhibitors for the proteinases A and B, and carboxypeptidase Y have been isolated from yeast (6,9-14) some of their properties are summarized in Table III. Two subgroups of inhibitors for the proteinases A and B, respectively, have been separated according to differences in electrophoretic mobility, i.e. in isoelectric point (see Table III). In early experiments (6), evidence for four different species of proteinase A-inhibitors had been obtained, but only the species 1 2 and 1 3 could be further purified and characterized (6,11). It has been shown recently (Biinning and Holzer in press) that in contrast to Baker s yeast, which contains the two inhibitors for proteinase B (10), in several strains of Saccharomyces cerevisiae only I 2 could be demonstrated and in Saccharomyces carlsbergensis, only 1 1. Single cell cultures from Baker s yeast contain 1 1 1 2 in the same concentration ratio of 1 3 as observed in preparations from commercial Baker s yeast. 

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