Carboxypeptidase time course

The effects of increasing concentrations of 8-OHQ-5SA, OP, and aa D on the activity of carboxypeptidase at a constant substrate concentration of 0.02 M CGP are shown in Fig. 2. (Vallee and Neurath, 1955.) Activity of the inhibited reaction was expressed as per cent of the proteolytic coefficient observed at zero inhibitor concentration. The conditions of preincubation are indicated. Recent and unpublished data indicate the time course of the inhibitory effects of these agents OP in concentrations of 1 X 10" M causes 90 % inhibition of the reaction in 60 minutes. 80 % of the inhibition occurs in the first 15 minutes (Fig. 3), Addition of 1 X 10" M zinc ions to the enzyme thus inhibited restores enzymatic activity, demonstrating the reversibility of inhibition (unpublished results). Since inhibition did not occur when chelating agents were first incubated with zinc, cupric, or ferrous ions to form the respective metal chelate, it appeared that the sites of chelation of these compounds are responsible for the observed inhibition. Inhibition is therefore not caused by any structural similarity between the inhibitors and the substrate. 

Fig. 3. Time course of 1,10-phenanthroline inhibition, 1 X 10 M OP resulting in 90% inhibition of carboxypeptidase in 60 minutes. Conditions of assay and per cent activity were calculated as in Fig, 2.

Fig. 12. Enzyme tryptophan (A) and substrate dansyl (B) fluorescence during the time course of zinc carboxypeptidase catalyzed hydrolysis of DNs-Gly-L-Phe. Equal volume solutions of substrate and of enzyme, both 2.5 x 10 M, in 1 M NaCl-0.02 M Tris, pH 7.5, 25°, were mixed and the fluorescence of either tryptophan (A) or dansyl (B) was measured as a function of time under stopped-flow conditions in parallel samples, as shown by the oscilloscope tracings. Excitation was at 285 nm. Scale sensitivities for (A) and (B) are 100 mV/div. The existence of the E S complex is signalled either by (A) the suppression of enzyme tryrptophan fluorescence (quenching by the dansyl group) or (B) enhancement of the substrate dansyl group fluorescence (energy transfer from enzyme trjqrtophan). Taken from Ref. (182) with permission...

The excess of pepstatin (peak A2) was removed on a column of Bio-Gel P2 (Fig.4). The composition and terminal analyses of A1 are given in Table V. Carboxyl-terminal residue was determined from a time-course of digestion with carboxypeptidase A. High-voltage paper electrophoresis at pH 6.5 and thin-layered chromatography showed A1 to contain one major basic peptide but, in both cases, very faint additional spots could be detected. 

Fig. VIII-5. Schematic diagram and time course of liberation of amino acids from the C-terminal region of TNP-clupeine YI by the alternate action of carboxypeptidases B and A [from Suzuki and Ando, 1972 (2)]...

When the Colonel was asked about teaching and research some time ago he replied "They are one and the same. When I do research, I get materials for my lectures, and when I teach I get ideas for my research. Many of us have also learned from the Colonel how much one can teach while seeming to merely inquire about the day s research progress. Certainly, his research provided the rest of us with material for our teaching, and in many areas. His work on boranes and on carboxypeptidase are the standard textbook treatments already. He used group theory and symmetry extensively in his research., and his Harvard course on Molecular Symmetry and Molecular Orbitals provided a perspective not available from any of the standard texts. Writing - of papers, of books, of... 

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