Proteases aspartyl

The HIV-1 protease, like other retroviral proteases, is a homodimeric aspartyl protease (see Fig. 1). The active site is formed at the dimer interface, with the two aspartic acids located at the base of the active site. The enzymatic mechanism is thought to be a classic acid-base catalysis involving a water molecule and what is called a push-pull mechanism. The water molecule is thought to transfer a proton to the dyad of the carboxyl groups of the aspartic acids, and then a proton from the dyad is transferred to the peptide bond that is being cleaved. In this mechanism, a tetrahedral intermediate transiently exists, which is nonconvalent and which is mimicked in most of the currently used FDA approved inhibitors. 

Partaledis JA, Yamaguchi K, Tisdale M, Blair EE, Falcione C, Maschera B, Myers RE, Pazhanisamy S, Futer O, CuHinan AB et al (1995) In vitro selection and characterization of human immunodeficiency virus type 1 (HIV-1) isolates with reduced sensitivity to hydrox-yethylamino sulfonamide inhibitors of HIV-1 aspartyl protease. J Virol 69 5228-5235 Patick AK (2006) Rhinovirus chemotherapy. Antiviral Res 71 391-396... 

Cathepsin D. The design of inhibitors of the aspartyl protease cathepsin D started from a virtual library of peptide analogs that contained the typical hydroxyethylamine isoster for the cleavable peptide bond. As the availability of starting materials would have generated a library of about 1 billion compounds, virtual screening was applied to reduce this multitude of candidate structures to a reasonable number. The backbone of a peptide... 

Plasmepsin II. The malarial aspartyl protease plasmepsin II has a significant homology (35%) to cathepsin D. Correspondingly, the very same approach as for the cathepsin D inhibitors (see above) was followed. The best inhibitors have Ki values of 2-10nM, a molecular weight <650, moderate selectivity vs. cathepsin D, the most closely related human protease, log P values <4.6, and no apparent binding to human serum albumin, for example, compound 36 Ki plasmepsin II = 2.0nM, Ki cathepsin D = 9.8nM Fig. 16.5) [111]. 

Haque TS, Skillman AG, Lee CE, Habashita H, Gluzman lY, Ewing TJA, Goldberg DE, Kuntz ID, Ellman JA. Potent, low-molecnlar-weight non-peptide inhibitors of malarial aspartyl protease plasmepsin II. J Med Chem 1999 42 1428-40. 

These must be worthwhile objectives and the recent identification by a number of research groups (see Skovronsky and Lee 2000 for description and details) of P-secretase as the membrane-bound aspartyl protease (RACE), S-site APP cleaving enzyme, paves the way for developing possible chemical inhibitors of its activity for experimental and clinical evaluation, although that remains for the future. 

The carboxyl proteases are so called because they have two catalytically essential aspartate residues. They were formerly called acid proteases because most of them are active at low pH. The best-known member of the family is pepsin, which has the distinction of being the first enzyme to be named (in 1825, by T. Schwann). Other members are chymosin (rennin) cathepsin D Rhizopus-pepsin (from Rhizopus chinensis) penicillinopepsin (from Penicillium janthinel-lum) the enzyme from Endothia parasitica and renin, which is involved in the regulation of blood pressure. These constitute a homologous family, and all have an Mr of about 35 000. The aspartyl proteases have been thrown into prominence by the discovery of a retroviral subfamily, including one from HIV that is the target of therapy for AIDS. These are homodimers of subunits of about 100 residues.156,157 All the aspartyl proteases contain the two essential aspartyl residues. Their reaction mechanism is the most obscure of all the proteases, and there are no simple chemical models for guidance. 

Figure 1.9 Reaction cycle for an aspartyl protease illustrating the conformational changes within the active site that attend enzyme turnover. Source Model based on experimental data summarized in Northrop (2001). 

Table 5.4 Hypothetical experiment measuring the IC50 values of a competitive inhibitor for HIV aspartyl protease and for human renin, at a fixed substrate concentration of 50 iM...

Example of Scheme C When Kf K, Aspartyl Protease Inhibitors... 

Figure 6.18 Structure of HIV-1 aspartyl protease in the flap open (left panel) and flap closed conformation with an active site-directed inhibitor bound right panel). See color insert.

The pepsin, activated by cleavage of a proenzyme, has two putative active site domains comprising hydrophobic-hydrophobic-Asp-Thr-Gly amino acids, is potentially glycosylated and has a free cysteine residue which may allow it to form dimers, as in the case of human and Plasmodium falciparum-derived aspartyl proteases (Longbottom et al., 1997). However,... 

The human retrovirus HIV can be controlled using chemotherapy directed at the reverse transcriptase and aspartyl protease encoded by the viral genome as with other microbial pathogens, however, resistance to drug therapy becomes a major problem. Figure 7.3 shows a crystal structure (PDB 1HXW) of the HIV protease, where mutated amino acids (shown in cyan) lead to disrupted binding of the clinically effective inhibitor ritonavir [24]. 

For example, with the crystal structure of the aspartyl protease from human immundeficiency virus (HIV-1) in 1989 came the opportunity to design molecules to block this important enzyme that acts as a molecular scissors. HIV is the virus responsible for AIDS. Essential to viral replication, the HIV protease cuts long strands composed of many proteins into the functional proteins found in mature virus particles. This proteolysis occurs at the very end of the HIV replication cycle (Figure 7-1). The three-dimensional structural information derived from the x-ray crystal structure, combined with computer modeling techniques, allowed chemists to design potent, selective inhibitors of the protease enzyme (Figure... 

Additional endoproteases may be shown to play a role in neuropeptide biosynthesis. Leading candidates are the mammalian homolog of the yeast aspartyl protease-3 (YAP-3) and a thiol protease. The processing of proANF, which involves cleavage after a single Arg residue in proANF, cannot involve PCI or PC2 since there are negligible amounts of these PCs in the heart. 

The functions of these protein and their interactions with each other in the complex and in y-secretase activity are not yet fully defined. PS1 may act as an aspartyl protease itself function as a cofactor critical for the activity of... 

Yan, R., Bienkowski, M. J., Shuck, M. E. et al. Membrane-anchored aspartyl protease with Alzheimer s disease P-secretase activity. Nature 402 533-537,1999. 

The life cycle of many viruses, including retroviruses, depends on viral proteases that cleave viral glycoproteins into individual polypeptides, and these enzymes are necessary for viral replication. NO can inactivate coxsackievirus [136]. Since cysteine proteases are critical for the virulence and replication of many viruses, nitrosation of viral cysteine proteases may be a mechanism of antiviral host defense. NO mediates nitrosation of cysteine and aspartyl proteases of HIV-1, and it was suggested that this... 

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