ABSTRACT
Purification of cathepsin Β from buffalo-spleen, a hitherto unstudied system has been achieved by a simple procedure developed by incorporating suitable modifications in the existing methods for isolation of the enzyme from other sources. The purified enzyme has a molecular weight of 25 KDa and its Stokes radius was found to be 2·24 nm. Effects of several reducing agents, urea and thiol-protease inhibitors such as leupeptin and antipain, have been studied and the data unequivocally support the contention that the buffaloenzyme is similar to cathepsin Β from other tissues with respect to these properties.
ABSTRACT
Some properties of a fragment of bovine serum albumin containing residues 184-582 of the protein sequence, produced by cyanogen bromide cleavage, have been reported. Urea-induced difference spectra of the fragment showed considerable exposure of aromatic chromophores by 8 Μ urea. Reversible unfolding of the fragment by urea, as followed by difference spectral measurements at 30°C, pH 7·0, occurred in two distinct steps involving at least 3 major conformational states, namely the native (N), intermediate (X) and completely denatured (D) states. The co-operativity values for the two transitions, N X and X Dwere found to be 4·0 and 16·4, respectively. Analysis of the data on bilirubin binding to bovine serum albumin and its fragment suggested that the fragment retains significant amount of its native structure. However, hydrodynamic parameters such as Stokes radius (3·14 nm), diffusion coefficient (6·98 × 10-7cm2/s) and frictional ratio (1·32) obtained by analytical gel chromatography as well as intrinsic viscosity (4·31 ml/g) indicates some asymmetry in the fragment molecule.