Different requirements for productive interaction between the active site of HIV-1 proteinase and substrates containing -hydrophobic*hydrophobic- or -aromatic*pro- cleavage sites.

The sequence requirements for HIV-1 proteinase catalyzed cleavage of oligopeptides containing two distinct types of junctions (-hydrophobic*hydrophobic- or -aromatic*Pro-) has been investigated. For the first type of junction (-hydrophobic*hydrophobic-) the optimal residues in the P2 and P2' positions were found to be Val and Glu, respectively, in accord with recent statistical analysis of natural cleavage sites [Poorman, R. A., Tomasselli, A. G., Heinrikson, R. L., & Kézdy, F. J. (1991) J. Biol. Chem. 266, 14554-14561]. For the -aromatic*Pro- type of junction, in the specific sequence context studied here, the value of Glu in the P2' position was again observed. An explanation for the inefficient cleavage observed for peptides with the sequence -Val-Tyr*Pro- has been provided from molecular modeling of the putative enzyme-substrate complex. A significant effect upon cleavage rates due to the amino acid in the P5 position has also been documented. While lysine in the P5 position in one sequence of the -hydrophobic*hydrophobic- type produces a peptide cleaved very efficiently (kcat greater than 15 s-1 for Lys-Ala-Arg-Val-Nle*p-nitrophenylalanine-P2'-Ala-Nle-NH2, for P2' = Glu, Gln, Ile, Val, or Ala), for substrates of the -aromatic*Pro- type, the P5 residue can exert either a positive or negative effect on cleavage rates. These results have again been interpreted in light of molecular modeling. We suggest that interaction of the substrate sequence on the periphery of the active site cleft may influence the match of the enzyme-substrate pair and, hence, control the efficiency of catalysis.(ABSTRACT TRUNCATED AT 250 WORDS)

[1]  F. Toma,et al.  A proton NMR investigation of proline-24 cis-trans isomerism in corticotropin 1-32 and related peptides. , 1978, Biochimica et biophysica acta.

[2]  Jones Ta,et al.  Diffraction methods for biological macromolecules. Interactive computer graphics: FRODO. , 1985, Methods in enzymology.

[3]  B. Dunn,et al.  A systematic series of synthetic chromophoric substrates for aspartic proteinases. , 1986, The Biochemical journal.

[4]  D. Baltimore,et al.  Standardized and simplified nomenclature for proteins common to all retroviruses , 1988, Journal of virology.

[5]  C. Debouck,et al.  Human immunodeficiency virus 1 protease expressed in Escherichia coli behaves as a dimeric aspartic protease. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[6]  C. Debouck,et al.  Peptide substrates and inhibitors of the HIV-1 protease. , 1989, Biochemical and biophysical research communications.

[7]  R. Blake,et al.  The Michaelis constants of a nonchromogenic substrate may be determined using a chromogenic substrate. , 1989, Archives of biochemistry and biophysics.

[8]  A Wlodawer,et al.  Structure of complex of synthetic HIV-1 protease with a substrate-based inhibitor at 2.3 A resolution. , 1989, Science.

[9]  M. Jaskólski,et al.  Conserved folding in retroviral proteases: crystal structure of a synthetic HIV-1 protease. , 1989, Science.

[10]  M. Navia,et al.  Three-dimensional structure of aspartyl protease from human immunodeficiency virus HIV-1 , 1989, Nature.

[11]  V. Kostka,et al.  Sub‐site preferences of the aspartic proteinase from the human immunodeficiency virus, HIV‐1 , 1990, FEBS letters.

[12]  C. Vlahos,et al.  Substitutions at the P2' site of gag p17-p24 affect cleavage efficiency by HIV-1 protease. , 1990, Biochemical and biophysical research communications.

[13]  I. Weber,et al.  Comparison of inhibitor binding in HIV‐1 protease and in non‐viral aspartic proteases: the role of the flap , 1990, FEBS letters.

[14]  A. Tomasselli,et al.  Ribonuclease A as a substrate of the protease from human immunodeficiency virus-1. , 1990, The Journal of biological chemistry.

[15]  A Wlodawer,et al.  X-ray crystallographic structure of a complex between a synthetic protease of human immunodeficiency virus 1 and a substrate-based hydroxyethylamine inhibitor. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[16]  I B Duncan,et al.  Rational design of peptide-based HIV proteinase inhibitors. , 1990, Science.

[17]  D. Norbeck,et al.  Design, activity, and 2.8 A crystal structure of a C2 symmetric inhibitor complexed to HIV-1 protease. , 1990, Science.

[18]  R. Dixon,et al.  Crystallographic analysis of a complex between human immunodeficiency virus type 1 protease and acetyl-pepstatin at 2.0-A resolution. , 1991, The Journal of biological chemistry.

[19]  W. Farmerie,et al.  Sensitive, soluble chromogenic substrates for HIV-1 proteinase. , 1990, The Journal of biological chemistry.

[20]  V. Kostka,et al.  Hydrolysis of synthetic chromogenic substrates by HIV-1 and HIV-2 proteinases. , 1990, Biochemical and biophysical research communications.

[21]  T. Copeland,et al.  Comparison of the HIV‐1 and HIV‐2 proteinases using oligopeptide substrates representing cleavage sites in Gag and Gag‐Pol polyproteins , 1991, FEBS letters.

[22]  I. Weber,et al.  Studies on the role of the S4 substrate binding site of HIV proteinases , 1991, FEBS letters.

[23]  A. Wlodawer,et al.  The complexities of AIDS : an assessment of the HIV protease as a therapeutic target , 1991 .

[24]  J. Kay,et al.  Mutating P2 and P1 residues at cleavage junctions in the HIV‐1 pol polyprotein Effects on hydrolysis by HIV‐1 proteinase , 1991, FEBS letters.

[25]  J. Springer,et al.  Structure and function of retroviral proteases. , 1991, Annual review of biophysics and biophysical chemistry.

[26]  A Wlodawer,et al.  Structure at 2.5-A resolution of chemically synthesized human immunodeficiency virus type 1 protease complexed with a hydroxyethylene-based inhibitor. , 1991, Biochemistry.

[27]  I. Weber,et al.  Comparative analysis of the sequences and structures of HIV‐1 and HIV‐2 proteases , 1991, Proteins.

[28]  A. Tomasselli,et al.  A cumulative specificity model for proteases from human immunodeficiency virus types 1 and 2, inferred from statistical analysis of an extended substrate data base. , 1991, The Journal of biological chemistry.

[29]  P E Wright,et al.  Defining solution conformations of small linear peptides. , 1991, Annual review of biophysics and biophysical chemistry.