Aspartic-type proteases

A similar situation exists in the acidic, aspartic-type proteases, where two acidic amino acid residues must interact to split the peptide bond in the substrate. These carboxylic groups also must dissociate differently, and therefore their pK values must be different. 

Seelmeier, S., Schmidt, H., Turk, V., and von der Helm, K.( 1988). Human immunodeficiency virus has an aspartic-type protease that can be inhibited by pepstatin A. Proc. Natl. Acad. Sci. U. S. A. 85,6612-6616. 

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... 

LaPointe CF, Taylor RK. 2000. The type 4 prepilin peptidases comprise a novel family of aspartic acid proteases. J. Biol. Chem. 275 1502-10... 

There are several types of proteases. HIV protease is an example of aspartic acid proteases. The structure of HIV protease as determined by X-ray crystallography is shown in Fig. 21.11. How the peptide bond is cleaved is shown schematically in Fig. 7.3. The set of two aspartic acid residues shown catalyzes the addition of water molecule to gag-pol protein (a). The result is the formation of an intermediate shown as (b) in the figure, (b) is known to decompose into two separate entities as shown in (c). This completes the hydrolysis. The region in the gag-pol protein where this splitting occurs has an amino acid sequence of -Leu-Asn-Phe-Pro-lle- (Asn=aparagine, He=isoleucine, Leu=leucine, Phe=phenyl alanine, Pro=proline). 

Based on their sequence homology, disulfide connectivity, and cysteine location within the sequence and chemistry of the reactive site. Pis can be assigned to distinct families, as classified by Laskowski and Kato. Kunitz-type, Bowman—Birk-type, Potato type I and type II, and squash inhibitors are members of these families shown in Table 3. For inhibitors not falling into these classifications more families have been proposed. Pis can also be classified by their target/mode of action. Plants have been found to express Pis that target serine proteinases, cysteine proteinases, aspartic proteinases, and metallo-proteinases. Serine and cysteine protease inhibitors are the best-studied PIs. ... 

Weihofen, A., Lemberg, M.K., Friedmann, E., et al. (2003) Targeting presenilin-type aspartic protease signal peptide peptidase with y-secretase inhibitors. J. Biol. Chem., 278, 16528-16533. 

A number of betaines of type (7) have been prepared as potential antibiotic analogues of fi-lactams and were found to inhibit serine and aspartic proteases <92EJM193,93JPP(45)466>. 

A characteristic feature of catalysis by the aspartic proteases is a tendency, with certain substrates, to catalyze transpeptidation reactions of the following type. 

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). 

Solanum (potato) Kunitz PEPs inhibit the aspartic protease cathepsin D as well as trypsin [125-134] and potato cysteine protease inhibitor (PCPI) inhibits a variety of cysteine proteases [185-188]. The crystal structures of soybean trypsin inhibitor (STI) [362, 368] and of Erythrina trypsin inhibitor (ETI) [350] have been determined. The structure of this type of plant Kunitz serine PIP involves a [3-barrel formed by 6 loop-linked antiparallel [3-strands with a lid formed by 6 further loop-linked antiparallel [3-strands. The scissile bond is located within a loop that extends out from the surface of the [3-barrel [350, 362, 368]. 

Figure 4 Chemical tools for the study of y-secretase. Transition-state anaiog inhibitors inciude hydroxyi-containing moieties that interact with the catalytic aspartates of aspartyl proteases. Helical peptides mimic the APR transmembrane domain and interact with the substrate docking site on the protease. These potent inhibitors were converted into affinity labeling reagents that contain a chemicaiiy reactive bromoacetyi or photoreactive benzophenone for covalent attachment to the protein target and a biotin moiety to allow isolation and detection of the labeled protein. Both types of chemical probes interacted with the two presenilin subunits but at distinct locations, which suggests that both the active site and the docking site of y-secretase lie at the interface between these subunits.

Weihofen A, Binns K, Lemberg MK, Ashman K, MartogUo B. Identification of signal peptide peptidase, a preseniUn-type aspartic protease. Science 2002 296 2215-2218. 

Aspartic proteases have also been proposed as the ECE of primary interest. Enzymes of this type have been isolated from a range of tissues and have been shown to cleave big ET-1 to ET-1. The case has been proposed for pepsin, cathepsin D and cathepsin E being the important enzyme, the latter being very attractive since the human enzyme has been shown to give only ET-1 and the C-terminal fragment (22-38) as big ET-1 breakdown products [62]. However, their physiological roles continue to be questioned as they require an acidic environment for activity. Inhibitors of the aspartic protease from rat lung tissue have been reported [63] but there is no information on inhibitors of the other aspartic enzymes. 

Loeb, D. D., Hutchison, C. A., Edgell, M. H., Farmerie, W. G., and Swanstrom, R.(1989). Mutational analysis of human immunodeficiency virus type 1 protease suggests functional homology with aspartic proteinases. y. Virol. 63, 111-121. 

The major driving force for the catalytic effect of serine proteases appeared to be associated with the electrostatic stabilisation of the (- + -) charge distribution, representing the Ser...His...Asp triad at the active site. This hypothesis is further supported by the experiments of Corey et al. (1992) who displaced the ionised aspartate in trypsin from the site 102 to the more distant site 214 by designing the D102S/S214E(D) double mutant. The rate reduction for two different substrates, as compared to the wild-type enzyme, was found to be 3 orders of magnitude less than for... 

Fig. 10.4 (continued) tide bond cleavage. In the cysteine (and also serine and threonine) proteases, the nucleophile is the protease type amino acid (in this case cysteine) which forms a covalent bond with the carbon atom of the bond to be cleaved (covalent catalysis) in contrast to the metalloprotei-nases and aspartic proteases which use an activated water molecule to attack the carbon atom to be cleaved (noncovalent catalysis). In covalent catalysis, a nearby histidine residue normally functions as a base to activate the mechanism, whereas in noncovalent catalysis, the protease type serves as an acid and base, with an ancillary histidine (aspartate proteases) or aspartate or glutamate residue acting as the nucleophile (Fig. 8.2b) (Modified from Fig. 9.18 in Berg., et al., Biochemistry, 5th Ed. 2002, W.H. Freeman Co., New York)...

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