On the purity of 3X-recrystallized bovine alpha-chymotrypsin

The results of an electrospray-mass spectrometric analytical study of aqueous solutions of fifteen commercial samples of 3X-recrystallized bovine alpha-chymotrypsin are presented and discussed. It was found that only six samples were predominantly alpha-chymotrypsin and that two samples contained no alpha-chymotrypsin at all. The remaining seven samples were found to be mixtures of alpha-chymotrypsin with other chymotrypsins and, in some cases, neochymotrypsinogens. The majority of the results are rationalised in terms of previously postulated and/or observed products of proteolytic activation of bovine chymotrypsinogen A. However, evidence is also presented for the presence in many of the samples of three new serine proteases, of significantly lower molecular masses than alpha-chymotrypsin, which cannot at present be explained. The paper is concluded with a brief discussion of the implications of the analytical findings for enzymological studies.