Enzymes aspartic proteases

There are multiple examples of drug action that depend on enzyme inhibition, including inhibitors of acetylcholinesterase, angiotensin converting enzyme, aspartate protease, carbonic anhydrase, cyclooxygenases, dihydrofolate reductase, DNA/RNA polymerases, monoamine oxidases, Na/K-ATPase, neuraminidase, and reverse transcriptase. 

Mammals, fungi, and higher plants produce a family of proteolytic enzymes known as aspartic proteases. These enzymes are active at acidic (or sometimes neutral) pH, and each possesses two aspartic acid residues at the active site. Aspartic proteases carry out a variety of functions (Table 16.3), including digestion pepsin and ehymosin), lysosomal protein degradation eathepsin D and E), and regulation of blood pressure renin is an aspartic protease involved in the production of an otensin, a hormone that stimulates smooth muscle contraction and reduces excretion of salts and fluid). The aspartic proteases display a variety of substrate specificities, but normally they are most active in the cleavage of peptide bonds between two hydrophobic amino acid residues. The preferred substrates of pepsin, for example, contain aromatic residues on both sides of the peptide bond to be cleaved. 

FIGURE 16.26 Acyl-enzyme and amino-enzyme intermediates originally proposed for aspartic proteases were modeled after the acyl-enzyme intermediate of the serine proteases. 

The first hint that two active-site carboxyl groups—one proto-nated and one ionized—might be involved in the catalytic activity of the aspartic proteases came from studies of the pH dependence of enzymatic activity. If an ionizable group in an enzyme active site is essential for activity, a plot of enzyme activity versus pH may look like one of the plots at right. 

It is worth noting here that inhibitors that interact with enzyme active site functionalities in ways that mimic the structure of covalent intermediates of catalysis can bind with very high affinity. This was seen in Chapter 1 with the example of statine-and hydroxyethylene-based inhibitors of aspartic proteases other examples of this inhibitor design strategy will be seen in subsequent chapters of this text. 

D. H. Rich, M. S. Bematowicz, N. S. Agarwal, M. Kawai, F. G. Salituro, and P. G. Schmidt, Inhibition of aspartic proteases by pepstatin and 3-methylstatine derivatives of pepstatin. Evidence for collected-substrate enzyme inhibition, Biochemistry 24 3165... 

Such an intermediate is known to be formed in reactions catalyzed by trypsin, chymotrypsin, thrombin, other enzymes of the blood-clotting cascade (except angiotensinconverting enzyme, which is an aspartic protease). An acyl-serine intermediate is also formed in the acetylcholinesterase reaction. The active site serine of this enzyme and the serine proteases can be alkylated by diisopropyl-fluorophosphate. See also Active Site Titration... 

Aspartate a-decarboxylase 753, 755 Aspartate p-decarboxylase 746 Aspartate racemase 741 Aspartic acid (Asp, D) 52, 53s biosynthesis 517 pXa value of 293, 487 Aspartic proteases 621-625 Aspartyl aminopeptidase 621 p-Aspartyl phosphate 539, 540s Assays of enzyme activity 456 Assembly core of virus shell 365 Assembly pathway... 

Proteases are enzymes that cleave proteins by hydrolyzing peptide bonds. On the basis of their catalytic mechanisms, they can be classified into five main types proteases that have an activated cysteine residue (cysteine proteases), an activated aspartate (aspartate proteases), a metal ion (metalloproteases), or an activated threonine (threonine proteases), and proteases with an active serine (serine proteases). Within each type, enzymes are separated into clans (also referred to as superfamilies ) based on evidence of evolutionary relationship [60, 61] from the linear order of... 

In general, hydrolytic enzymes can be classified based on the type of reaction catalyzed, the nature of the enzyme active site, and/or evolutionary relationships among enzymes, as derived from primary sequence data. Among proteases, gross functional distinctions are made between serine proteases, aspartic proteases, cysteine proteases, and metalloproteases. Each of these groups includes a diverse range of enzymes of distinctive size and structure for example, an aminopeptidase isolated from B. lichenformis was found to have a molecular mass of 34,000, whereas an E. coli aminopeptidase had a mass of 400,000 (Rao et al., 1998). 

Several pharmaceutical enzymes belong to the group of serine-histidine estero-proteolytic enzymes (serine proteases), which display their catalytic activity with the aid of an especially reactive serine residue, whoso p-hydroxyi group forms a covalent bond with the substrate molecule. This reaction takes place by cooperation with the imidazole base of histidine. The specificity of the enzymes is achieved by the characteristic strocture of their substrate-binding centers, which in these proteases are built according to the same principle. They consist of a hydrophobic slit formed by apolar aide chains of amino acids and a dissociated side chain-located carboxyl group of an aspartic add residue at the bottom. 

The pharmacological evidence compiled for y-secretase is indicative of the activity of an aspartic protease requiring at least one additional cofactor. The location of the active site within the membrane makes y-secretase quite unique. Currently, there is only one precedent for a similar, tricky enzyme, signal peptidase, which shares several features and most of the problems associated with inner-membrane location [25]. It will not, unfortunately, be easy to isolate and purify the membrane-stabilized protease while retaining its activity. It has, therefore, so far escaped crystallization and X-ray structure determination. Mutation analysis of the two conserved aspartic acids of all presenilins supports their key role in y-secretase... 

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