Catalytic aspartic acids

Fig. 1 A ribbon diagram of the crystal structure of a substrate complex of the homo-dimer HIV-1 protease (lkj7) (Prabu-Jeyabalan et al. 2002), Each monomer is shown in cyan and pink the substrate is shown in green, and the catalytic aspartic acids are highlighted in yellow...

The three-dimensional structures of human (left) and mouse renins (right) showing oligopeptide inhibitors bound in the active site cleft. The cleft lies between the N- and C-terminal domains of the enzyme and is approximately perpendicular to the plane of the page. It can accommodate 9-10 residues with the substrate/inhibitor bound in an extended conformation. The catalytic aspartic acid residues (not shown) are centrally placed at the base of the cleft. 

Mooser G, Hefta SA, Paxton RJ, Shively JE, Lee TD (1991) Isolation and sequence of an active-site peptide containing a catalytic aspartic acid from two Streptococcus sobrinus alpha-glucosyltransferases. J Biol Chem 266 8916-8922... 

The mechanism of the PTP hydrolysis reaction has two steps. First, phosphate is transferred from tyrosine to the cysteine residue of the P-loop, which generates a phosphoenzyme intermediate with concomitant release of tyrosine. This process is followed by hydrolysis of the phosphoenzyme to free enzyme and inorganic phosphate. Two active site residues are of primary importance during the catalytic cycle the nucleophilic cysteine of the P-loop and an aspartate on a nearby flexible loop, which serves as a general acid/base catalyst (Fig. 3). After attack of the cysteine on phosphotyrosine, tyrosine can be expelled as the protonated phenol after proton donation by the catalytic aspartic acid, which forms the phosphoenzyme intermediate and free tyrosine. The aspartate anion then deprotonates the hydrolytic water molecule that attacks phosphocysteine, which liberates inorganic phosphate (Fig. 4) (9). 

Figure 2. Stereoview of a renin inhibitor, Smo-Phe-Atm-ACDMH (yellow), bound in the renin model binding site (green, catalytic aspartic acids in red).

Figure 5. Stereoview of the X-ray crystal structure of a renin inhibitor, Boc-Phe(CHOHCH2)Gly-ACHPA-LEU-AMPMA (yellow), bound in endothiapepsin (cyan, catalytic aspartic acids in red) [17], A hydrogen bond between the hydroxyl oxygen and the Thr219(NH) (magenta) is shown by a white dotted line.

Figure 7. Stereoview of the human renin X-ray crystal structure bound with an inhibitor [43]. The inhibitor is shown in yellow and the enzyme in red with the catalytic aspartic acids, the polyproline loop and Prolll on helix his,2 colored cyan.

Figure 11. Stereoview of the bound conformation of 29 (black lines) resulting from an AUTODOCK simulation with HIV-1 protease [72], Asp25/Aspl25 (the catalytic aspartic acids), Ile50/Ilel50 and ArglOS are also shown (gray lines).

HIV-1 PR is a symmetrical homodimer consisting of two identical subunits of 99 amino acids. Its active site is formed at the dimer interface and contains two conserved, catalytic aspartic acid residues. A water molecule bound to the enzyme between the two aspartates acts as the nucleophile for catalysis. Each monomer contains a prominent (3-hairpin loop, known as the flap, that projects over the substrate-binding cleft. These flaps are highly flexible and can undergo large localized conformational changes upon substrate and inhibitor binding [25-27]. 

A central hydroxyl or diol group which binds to the catalytic aspartic acid carboxyls. 

Hybrid potentials have been used to understand the mechanism of the human immunodeficiency virus (HIV) protease with the ultimate aim of being able to help in the design of inhibitors which could be useful as AIDS therapies. This enzyme, which catalyzes the hydrolysis of peptide bonds, is a homodimer. Its active center is at the interface of the two chains and consists of two catalytic aspartic acid residues from identical positions in each of the two chains. Although the aspartates are equivalent in the sequence, they are not equivalent when the substrate is present. It is known that when the enzyme is active one of the aspartic residues is protonated and it is thought that there is a lytic water molecule that is also involved in the catalysis. 

The continued derivation now shifts to the more specific problem of a ligand binding to wild type HIV-1 protease and its mutants. In this case P will be now be called and P will be called P,, y, where WT and MU stand for wild type and mutant protease respectively. HIV-1 protease is composed of two noncovalently associated structurally identical monomers, the active site contains two conserved catalytic aspartic acid residues, one from each monomer. Furthermore, the active site is C2 symmetric. Any active site mutation will result in structural effects in two different locations, one on each monomer, therefore, mutation does not break the C2 symmetry. As a result of these considerations, we can assume that the symmetry numbers for P,. and P are the same. This will also hold for LP, and LP. The ratio of symmetry numbers will then be equal to unity and the natural logarithm then zeros out the first term of equation (15). 

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