Procarboxypeptidases, protein

This zinc-dependent enzyme [EC 3.4.17.1], a member of the peptidase family M14, catalyzes the hydrolysis of peptide bonds at the C-terminus of polypeptides. Little hydrolytic action occurs if the C-terminal amino acid is aspartate, glutamate, arginine, lysine, or proline. Car-boxypeptidase A is formed from a precursor protein, procarboxypeptidase A. 

The inactive precursors are called trypsinogen, pepsinogen, chymotrypsino-gen, and procarboxypeptidase. These precursors are converted to the active enzymes by hydrolytic cleavage of a few specific peptide bonds under the influence of other enzymes (trypsin, for example, converts chymotrypsinogen to chymotrypsin). The digestive enzymes do not appear to self-destruct, probably because they are so constructed that it is sterically impossible to fit a part of one enzyme molecule into the active site of another. In this connection, it is significant that chymotrypsin attacks denatured proteins more rapidly than natural proteins with their compact structures of precisely folded chains. 

Zymogens have been considered to be inactive precursors of enzymes and the activation process to involve the generation of a catalytic or substrate binding site or both 44). Recently, Behnke and Vallee (50) found that the spectral properties of cobalt-substituted procarboxypeptidase A closely resemble those of the cobalt enzyme. Since these spectra were believed to be peculiar to enzymatically active proteins (5i), they investigated the intrinsic catalytic activity of the cobalt zymogen. Remarkably, with certain substrates, cobalt procarboxypeptidase was found to have as much activity, and in some cases even more than the native enzyme. These observations, as well as those of others (52), have questioned the entire concept of zymogens as inactive enzyme precursors. 

Fig. 1. Chromatography of bovine pancreatic juice on DEAE-cellulose (anionic proteins) and Amberlite IRC-50 (cationic proteins) (1). RNAase, ribonuclease ChTg-a, chymotrypsinogen A Tg, trypsinogen ProCp-B and Cp-B, procarboxypeptidase B and carboxypeptidase B DNAase, deoxyribonuclease ProCp-A, procarboxypeptidase A.

For preparative purposes, it is perhaps more convenient to start from pancreas acetone powder (pancreatin). Ninty-five per cent pure bovine procarboxypeptidase A has been obtained by ammonium sulfate fractionation of pancreatin extracts and isoelectric precipitations (47). When the proteins precipitated by 0.39 saturated ammonium sulfate are chromatographed on DEAE-cellulose in a concentration gradient of pH 8.0 phosphate buffer, the two last and most acidic peaks contain procarboxypeptidase A in an electrophoretically homogeneous form (48). The molecular weight of the protein determined by light scattering and sedimentation-diffusion is 94-96,000. Its isoelectric point in univalent buffers of 0.2 ionic strength is below 4.5. 

It is a proteolytic enzyme, present in the intestine in its inactive form (zymogen), trypsinogen. Trypsinogen is converted into its active form, trypsin, by enteropeptidase, a specialized proteolytic enzyme secreted by intestinal cells. Some free trypsin formed also catalyses the conversion of trypsinogen into trypsin. Trypsin can also convert chymotrypsinogen and procarboxypeptidase into chymotrypsin and carboxypeptidase, respectively. Trypsin has different amino acid specificity when compared with other proteolytic enzymes. Trypsin hydrolyses those peptide bonds whose carboxyl groups are contributed by Lys or Arg residues and if the next residue is not proline. The number of smaller peptides resulting from trypsin action is equal to the total number of Arg and Lys residues in the protein plus one. 

V. Villegas, J. C. Martinez, F. X. Aviles et al. Structure of the transition state in the folding process of human procarboxypeptidase A2 activation domain. Journal of Molecular Biology, 283 (1998), 1027 F. Chiti, N. Taddei, P. M. White et al. Mutational analysis of acylphosphatase suggests the importance of topology and contact order in protein folding. Nature Structural Biology, 6 (1999), 1005. 

The pancreatic peptidases are also secreted as zymogens (trypsinogen, chymotrypsinogen, proelastase, and procarboxypeptidase). They are synthesized in cells in the pancreas, where they are packaged into granules with a lipid-protein "coat." The granules accumulate at the apex of the cell and are secreted into a duct leading to the intestine. 

The zymogen trypsinogen is cleaved to form trypsin by enteropeptidase (a protease, formerly called enterokinase) secreted by the brush-border cells of the small intestine. Trypsin catalyzes the cleavages that convert chymotrypsinogen to the active enzyme chymotrypsin, proelastase to elastase, and the procarboxypeptidases to the car-boxypeptidases. Thus, trypsin plays a central role in digestion because it both cleaves dietary proteins and activates other digestive proteases produced by the panaeas. 

Propnrteins inactive protein precirtsots, which are activated by the removal (a highly specific reaction) of a peptide sequence. Exarrtples are various Secretory enzymes (see) (procarboxypeptidase, proelas-tase, prothrombin, etc.) hormone precursors (proinsulin, proparathormone, etc.), peptide toxins (promelli-tin, etc.) and Zymogens (see). 

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