Document 0789
 DOCN  M9440789
 TI    Human immunodeficiency virus type-1 reverse transcriptase and
       ribonuclease H as substrates of the viral protease.
 DT    9404
 AU    Tomasselli AG; Sarcich JL; Barrett LJ; Reardon IM; Howe WJ; Evans DB;
       Sharma SK; Heinrikson RL; Biochemistry Unit, Upjohn Laboratories,
       Kalamazoo, Michigan; 49001.
 SO    Protein Sci. 1993 Dec;2(12):2167-76. Unique Identifier : AIDSLINE
       MED/94129399
 AB    A study has been made of the susceptibility of recombinant constructs of
       reverse transcriptase (RT) and ribonuclease H (RNase H) from human
       immunodeficiency virus type 1 (HIV-1) to digestion by the HIV-1
       protease. At neutral pH, the protease attacks a single peptide bond,
       Phe440-Tyr441, in one of the protomers of the folded, active RT/RNase H
       (p66/p66) homodimer to give a stable, active heterodimer (p66/p51) that
       is resistant to further hydrolysis (Chattopadhyay, D., et al., 1992, J.
       Biol. Chem. 267, 14227-14232). The COOH-terminal p15 fragment released
       in the process, however, is rapidly degraded by the protease by cleavage
       at Tyr483-Leu484 and Tyr532-Leu533. In marked contrast to this p15
       segment, both p66/p51 and a folded RNase H construct are stable to
       breakdown by the protease at neutral pH. It is only at pH values around
       4 that these latter proteins appear to unfold and, under these
       conditions, the heterodimer undergoes extensive proteolysis. RNase H is
       also hydrolyzed at low pH, but cleavage takes place primarily at
       Gly436-Ala437 and at Phe440-Tyr441, and only much more slowly at
       residues 483, 494, and 532. This observation can be reconciled by
       inspection of crystallographic models of RNase H, which show that
       residues 483, 494, and 532 are relatively inaccessible in comparison to
       Gly436 and Phe440. Our results fit a model in which the p66/p66
       homodimer exists in a conformation that mirrors that of the heterodimer,
       but with a p15 segment on one of the protomers that is structurally
       disordered to the extent that all of its potential HIV protease cleavage
       sites are accessible for hydrolysis.
 DE    Amino Acid Sequence  Comparative Study  Hydrogen-Ion Concentration
       Hydrolysis  HIV Protease/*METABOLISM  HIV-1/*ENZYMOLOGY  Models,
       Molecular  Molecular Sequence Data  Protein Conformation  Protein
       Denaturation  Recombinant Proteins/METABOLISM  Reverse
       Transcriptase/*METABOLISM  Ribonuclease H, Calf Thymus/*METABOLISM
       Sequence Homology, Amino Acid  Substrate Specificity  JOURNAL ARTICLE

       SOURCE: National Library of Medicine.  NOTICE: This material may be
       protected by Copyright Law (Title 17, U.S.Code).