Biosynthesis of fatty acids and phenols by stationary-phase cultures of Aspergillus fumigatus.

With this method it was shown that the covalent binding of trypsin to CNBr-activated Sephadex stabilized the protein against the denaturing effect of concentrated urea solutions, in contradistinction to the similar agarose-bound trypsin, which behaved as the free trypsin. This agrees with activity measurements (Gabel et al., 1970), the latter derivative being inactive in 8M-urea, as is the free enzyme, whereas Sephadex-bound trypsin retains most of its activity even after long storage in urea solutions. The stabilization of the active surface of trypsin by Ca , found for the free enzyme (Sipos & Merkel, 1970), was not observed with the Sephadex-bound enzyme. The protein in solution unfolds at 60°C in the presence of Ca2+, as reflected by a red shift of the tryptophan fluorescence, whereas the fluorescence properties of Sephadex-bound trypsin did not change up to 70°C. At that temperature the fluorescence intensity increased, but no red shift was observable. This increase coincided with the loss of activity. It is suggested that the temperature-induced conformational transitions that inactivate trypsin in the absence and in the presence of Ca2+, as well as the Ca2+_ dependent transition leading from native trypsin to a more compact structure (Sipos & Merkel, 1970), are not possible in Sephadex-bound trypsin owing to a more-or-less extensive cross-linking between the protein and the rigid carrier. Attempts were made to identify those c-amino groups that contribute to the stabilization of Sephadex-bound trypsin against denaturing influences.