Peptide bond enzyme-catalyzed hydrolysis

Section 25.10 Selective hydrolysis can be accomplished by using enzymes to catalyze cleavage at specific peptide bonds. Carboxypeptidase-catalyzed hydrolysis can be used to identify the C-terminal amino acid. The N taminus is determined by chemical means. One reagent used for this purpose is SangCT s reagent, l-fluoro-2,4-dinitro-... 

Protease inhibitor (Section 28.13) A substance that interferes with enzyme-catalyzed hydrolysis of peptide bonds. 

The enzyme-catalyzed hydrolysis of a particular peptide bond is determined by two major factors, the susceptibility of that bond to the specific proteinase and the flexibility of the protein chain in the region of the bond. Thus a globular protein undergoes frequent fluctuations, and the sites of the peptide chains of highest mobility are the most susceptible to proteolytic reactions. On the other site, the... 

A group of proteolytic enzymes, including trypsin and chymotrypsin, can be used to catalyze the hydrolysis of specific peptide bonds. Trypsin catalyzes the hydrolysis of peptide bonds formed by the carboxyl groups of Arg and Lys chymotrypsin catalyzes the hydrolysis of peptide bonds formed by the carboxyl groups of Phe, Tyr, and Trp. 

Enzyme Catalyzes Hydrolysis of Peptide Bond Formed by Carboxyl Group of... 

Enzyme-Catalyzed Hydrolysis of Peptides (Amide Bonds)... 

If the enzyme-catalyzed hydrolysis of peptide bond involves a simple reversible reaction as shown by Equation 2.5 then, indeed, the enzyme must catalyze the rate of formation of peptide bond from amino acids (i.e., lq,-step), provided the amino acids do not react irreversibly with the enzyme. Incidentally, if the function of serine proteases is to catalyze both the rate of hydrolytic cleavage and the rate of formation of protein peptide bond, then, probably, these enzymes cannot digest the proteins that we eat and, consequently, the results would have been disastrous for all protein-eating creatures — which certainly Nature will never allow. Although the mechanisms of most of the enzyme-catalyzed reactions are unknown, even at a very rudimentary level, the mechanism of a-chymotrypsin-catalyzed hydrolysis of peptide bond has been relatively well understood. The reaction has been almost ascertained to involve acylation and deacylation of enzyme as shown by Equation 2.6. Widely accepted mechanisms for acylation and deacylation steps are shown in Scheme 2.6 and Scheme 2.7. ... 

Gaertner et al. [48] investigated peptide bond formation catalyzed by PEG-proteases in benzene. The substrate specificity of PEG-chymotrypsin for ester hydrolytic activity in aqueous solution was almost the same as that of unmodified enzyme [51] both PEG-modified and unmodified chymotrypsins catalyzed the hydrolysis of aromatic amino acid esters but these enzymes did not hydrolyze the basic amino acid ester. In the case of dipeptide synthesis in benzene, however, not only Bz-iyr-Phe-NHa but also Bz-Lys-Phe-NH2 was synthesized from corresponding amino acid derivatives by PEG-chymotrypsin. The change in the substrate specificity, which has been reported in a few cases, might be due to a surrounding environment on the enzyme molecule. 

Knowing how the protein chain is folded is a key ingredient m understanding the mechanism by which an enzyme catalyzes a reaction Take carboxypeptidase A for exam pie This enzyme catalyzes the hydrolysis of the peptide bond at the C terminus It is... 

Selective hydrolysis can be accomplished by using enzymes to catalyze cleavage at specific peptide bonds... 

Chymotrypsin (Section 27 10) A digestive enzyme that cat alyzes the hydrolysis of proteins Chymotrypsin selectively catalyzes the cleavage of the peptide bond between the car boxyl group of phenylalanine tyrosine or tryptophan and some other ammo acid... 

Carboxypeptidases are zinc-containing enzymes that catalyze the hydrolysis of polypeptides at the C-terminal peptide bond. The bovine enzyme form A is a monomeric protein comprising 307 amino acid residues. The structure was determined in the laboratory of William Lipscomb, Harvard University, in 1970 and later refined to 1.5 A resolution. Biochemical and x-ray studies have shown that the zinc atom is essential for catalysis by binding to the carbonyl oxygen of the substrate. This binding weakens the C =0 bond by... 

The most recent advance in treating HIV infections has been to simultaneously attack the virus on a second front using a protease inhibitor. Recall from Section 27.10 that proteases are enzymes that catalyze the hydrolysis of proteins at specific points. When HIV uses a cell s DNA to synthesize its own proteins, the initial product is a long polypeptide that contains several different proteins joined together. To be useful, the individual proteins must be separated from the aggregate by protease-catalyzed hydrolysis of peptide bonds. Protease inhibitors prevent this hydrolysis and, in combination with reverse transcriptase inhibitors, slow the reproduction of HIV. Dramatic reductions in the viral load in HIV-infected patients have been achieved with this approach. 

Different enzymes have different specificities. Some, such as amylase, are specific for a single substrate, but others operate on a range of substrates. Papain, for instance, a globular protein of 212 amino acids isolated from papaya fruit, catalyzes the hydrolysis of many kinds of peptide bonds. In fact, it s this ability to hydrolyze peptide bonds that makes papain useful as a meat tenderizer and a cleaner for contact lenses. 

Staphylococcal nuclease (SNase) is a single-peptide chain enzyme consisting of 149 amino acid residues. It catalyzes the hydrolysis of both DNA and RNA at the 5 position of the phosphodiester bond, yielding a free 5 -hydroxyl group and a 3 -phosphate monoester... 

Other interesting examples of proteases that exhibit promiscuous behavior are proline dipeptidase from Alteromonas sp. JD6.5, whose original activity is to cleave a dipeptide bond with a prolyl residue at the carboxy terminus [121, 122] and aminopeptidase P (AMPP) from E. coli, which is a prohne-specific peptidase that catalyzes the hydrolysis of N-terminal peptide bonds containing a proline residue [123, 124]. Both enzymes exhibit phosphotriesterase activity. This means that they are capable of catalyzing the reaction that does not exist in nature. It is of particular importance, since they can hydrolyze unnatural substrates - triesters of phosphoric acid and diesters of phosphonic acids - such as organophosphorus pesticides or organophosphoms warfare agents (Scheme 5.25) [125]. 

In mammalian cells, the two most common forms of covalent modification are partial proteolysis and ph osphorylation. Because cells lack the ability to reunite the two portions of a protein produced by hydrolysis of a peptide bond, proteolysis constitutes an irreversible modification. By contrast, phosphorylation is a reversible modification process. The phosphorylation of proteins on seryl, threonyl, or tyrosyl residues, catalyzed by protein kinases, is thermodynamically spontaneous. Equally spontaneous is the hydrolytic removal of these phosphoryl groups by enzymes called protein phosphatases. 

There are two main classes of proteolytic digestive enzymes (proteases), with different specificities for the amino acids forming the peptide bond to be hydrolyzed. Endopeptidases hydrolyze peptide bonds between specific amino acids throughout the molecule. They are the first enzymes to act, yielding a larger number of smaller fragments, eg, pepsin in the gastric juice and trypsin, chymotrypsin, and elastase secreted into the small intestine by the pancreas. Exopeptidases catalyze the hydrolysis of peptide bonds, one at a time, fi"om the ends of polypeptides. Carboxypeptidases, secreted in the pancreatic juice, release amino acids from rhe free carboxyl terminal, and aminopeptidases, secreted by the intestinal mucosal cells, release amino acids from the amino terminal. Dipeptides, which are not substrates for exopeptidases, are hydrolyzed in the brush border of intestinal mucosal cells by dipeptidases. 

Many enzymes have absolute specificity for a substrate and will not attack the molecules with common structural features. The enzyme aspartase, found in many plants and bacteria, is such an enzyme [57], It catalyzes the formation of L-aspartate by reversible addition of ammonia to the double bond of fumaric acid. Aspartase, however, does not take part in the addition of ammonia to any other unsaturated acid requiring specific optical and geometrical characteristics. At the other end of the spectrum are enzymes which do not have specificity for a given substrate and act on many molecules with similar structural characteristics. A good example is the enzyme chymotrypsin, which catalyzes hydrolysis of many different peptides or polypeptides as well as amides and esters. 

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