HIV protease substrates

Recently, two moderately active (V-hydroxyamide) analogues of the HIV protease substrate have been prepared in two different ways, with O-benzyl protection of the TV-hydroxy-a-amino acid residue.150 Boc-Phet >[CO-N(OH)]Gly-Ile-Phe-OMe is constructed by the mixed anhydride procedure in a step-by-step elongation of the peptide backbone. In contrast, Boc-Phei i[CO-N(OH)]Phe-Ile-Phe-OMe 49 is prepared by fragment condensation (Scheme 15). 

Scheme 15 Synthesis of an (TV-Hydroxvamide) Analogue of a HIV Protease Substrate by Fragment...

The syntheses of only a very few (A-aminoamide) peptides, with no examples of solid-phase synthesis, have been reported in the literature. The low yield in -coupling calls for the fragment condensation methodology, which has been applied to the synthesis of the HIV protease substrate analogues 79 and 80 (Scheme 23)J50]... 

Wang, G.T. et al. 1990. Design and synthesis of new fluorogenic HIV protease substrates based on resonance energy transfer. Tetrahedron Lett. 31, 6493-6496. 

There are other examples of HIV protease substrates that utilize the fluorescence energy transfer technique. Perhaps the strongest characteristic of these assays is that they provide a continuous readout of enzyme activity. Another advantage is their sensitivity (they use small concentrations of enzyme). The disadvantage of these assays is that they are susceptible to interference by some compounds (inhibition artifacts) because of inner and outer filter effects (see Notes 8-10). Other assays, for example, those based on radioactivity or high-performance liquid chromatography (HPLC) analysis of products are tedious to run, but are less susceptible to interference or inhibition artifacts. 

Table I. Kinetic Parameters for Fluorogenic HIV Protease Substrates ...

FIGURE 16.30 A mechanism for the incorporation of from H9 0 into peptide substrates in the HIV protease reaction. (Adapted from Hyland, ., et ai, 1991. BiochemisU y 30 8441-8453.)... 

Chellappan S, Kiran Kumar Reddy GS, All A, Nalam MN, Anjum SG, Cao H, Kairys V, Fernandes MX, Altman MD, Tidor B, Rana TM, Schiffer CA, Gilson MK (2007) Design of mutation-resistant HIV protease inhibitors with the substrate envelope hypothesis. Chem Biol Drug Des 69 455... 

A. Gupta, Y. Zhang, J.D. Unadkat, and Q. Mao. HIV protease inhibitors are inhibitors but not substrates of the human breast cancer resistance protein (BCRP/ABCG2). J Pharmacol Exp Ther. 310 334-341 (2004). 

Selected entries from Methods in Enzymology [vol, page(s)] Active site, structure, 241, 214 catalytic mechanism, 241, 223-224 crystal structure, 241, 214, 216 comparative studies with HIV protease [catalytic properties, 241, 205-224 evolutionary relationships, 241, 196-197 screening for HIV-1 protease inhibitors, 241, 318-321 structure, 241, 254-257, 280 substrate specificity, 241, 255, 283]. 

Figure 6. An example of inter-family target hopping between human and viral aspartyl proteases. The aspartyl protease active site is located at a homodimer interface in HIV and within a single domain in Cathepsin D, so sequence and structure alignments between these proteins cannot be constructed. By using an approach independent of sequence or structure homology to directly align the sites, SiteSorter finds that the HIV protease and Cathepsin D substrate sites are highly similar (identical chemical groups within 1 A are colored dark blue). It has been verified experimentally that Cathepsin D is susceptible to inhibition by HIV-protease inhibitors. ...

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