Carboxypeptidase surface

Carbon atom, 4. See also Atomic orbitals Carbon dioxide hydration, 197-199. See also Carbonic anhydrase Carbonic anhydrase, 197-199,200 Carbonium ion transition state, 154, 159 Carboxypeptidase A, 204-205 Catalysis, general acid, 153,164,169 in carboxypeptidase A, 204-205 free energy surfaces for, 160, 161 in lysozyme, 154... 

There is a third region of a protein that is neither on the surface nor in the interior but that is in a cleft. Such regions are often associated with enzyme action and examples show they have (1) intermediate mobility (e.g., tryptophan 62 of lysozyme or the tyrosine of carboxypeptidase) and (2) unfavorable energetics of exposed groups—the entatic state. 

Proteases of L. bulgaricus and L. helveticus contribute to the ripening of Swiss cheese (Langsrud and Reinbold 1973). Strains of thermo-duric lactobacilli are generally more proteolytic than S. thermophilus (Dyachenko et al. 1970). The proteinase activity of L. bulgaricus is optimal at pH 5.2-5.8 and is associated with the cell envelope (Argyle et al. 1976). Some strains of L. brevis (Dacre 1953) andL. lactis (Bottazzi 1962) are also proteolytic. Surface-bound aminopeptidase from L. lactis, characterized by Eggiman and Bachman (1980), is activated by cobalt and zinc ions and has optimum activity at pH 6.2-7.2. A surface-bound proteinase and carboxypeptidase are also present in L. lactis. 

The considerable detail to which we now can understand enzyme catalysis is well illustrated by what is known about the action of carboxypeptidase A. This enzyme (Section 25-7B and Table 25-3) is one of the digestive enzymes of the pancreas that specifically hydrolyze peptide bonds at the C-terminal end. Both the amino-acid sequence and the three-dimensional structure of carboxypeptidase A are known. The enzyme is a single chain of 307 amino-acid residues. The chain has regions where it is associated as an a helix and others where it is associated as a /3-pIeated sheet. The prosthetic group is a zinc ion bound to three specific amino acids and one water molecule near the surface of the molecule. The amino acids bound to zinc are His 69, His 196, and Glu 72 the numbering refers to the position of the amino acid along the chain, with the amino acid at the /V-terminus being number l. The zinc ion is essential for the activity of the enzyme and is implicated, therefore, as part of the active site. 

Figure 25-19 Steps in a possible mechanism of carboxypeptidase action, (a) The substrate is shown complexed to the enzyme surface through X, Y, Z, and W X is a nonpolar pocket Y is a hydrogen bond, possibly from OH of Tyr 248 Z is the prosthetic group, Zn and W is an...

The interdependence of protein surface and hydration of Sephadex has implications for confusion of the molecular mass experiments. The acid carboxypeptidase from A. saitoi is a glycoprotein, containing approximately 30% carbohydrate by weight [81, 82], Furthermore, a single protein band with molecular mass of 72,000 was detected in SDS-PAGE [79], This indicates that the carboxypeptidase is a homologous dimmer [79],... 

Zinc usually binds to proteins via residues of cysteine and histidine. Sometimes zinc is bound to residues of glutamate or aspartate. The zinc ion sometimes plays a catalytic role and sornetimes a structural role. In the latter case, it helps maintain the three-dimensional structure or conformation of the protein. For example carboxypeptidase A contains twr> atoms of zinc. One is required for catalytic activity and is bound to cysteine and histidine. The other, which plays a structural role, is bound only hr cysteine. Cytoplasmic supeioxide dismutase is a dimer, ft contains one atom of Cu and one of Zn + per subunit. The zinc is bound via three residues of histidine and one residue of aspartate. It is buried deep within the enzyme and serves a structural role. The copper atom is bound via four residues of histidine. It resides dose to the surface of the protein and participates in the chemistry of catalysis. 

Carboxypeptidase A is monomeric (Mj 34 500) and exists in three forms (a, P and y) which contain 307, 305 and 300 amino acids respectively. Near the surface of the protein lies a pocket in which a Zn ion is bound to the protein backbone by one didentate Glu and two His residues. A 5-coordinate coordination sphere is completed by a water molecule (Figure 28.24a). 

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