Document 0458
 DOCN  M9550458
 TI    Effect of pH and nonphysiological salt concentrations on human
       immunodeficiency virus-1 protease dimerization.
 DT    9505
 AU    Tyagi SC; Simon SR; Carter CA; Department of Medicine, University of
       Missouri, Columbia 65212.
 SO    Biochem Cell Biol. 1994 May-Jun;72(5-6):175-81. Unique Identifier :
       AIDSLINE MED/95142975
 AB    Human immunodeficiency virus-1 (HIV-1) protease is catalytically active
       as a dimer of identical subunits that associate through noncovalent
       interactions. To investigate the forces stabilizing HIV-1 protease in
       its active form, we have studied the effects of pH and salts on
       structure and function of the enzyme. Enzymatic activity was measured by
       following the hydrolysis of a fluorogenic substrate. Dissociation of the
       dimer into its subunits was monitored by gel filtration, while
       conformational changes in the enzyme were probed by measurements of
       intrinsic tryptophan fluorescence. Mg2+ ions were capable of
       dissociating the dimeric enzyme with a concomitant red shift and
       increase in quantum yield of the tryptophan fluorescence, indicating
       increased accessibility of tryptophan to the aqueous environment. These
       structural changes also were associated with a loss of catalytic
       activity which was insensitive to substrate concentration, consistent
       with noncompetitive inhibition. Both structural and functional changes
       could be attributed to binding of Mg2+ ions to a site with an apparent
       dissociation constant of approximately 2 M. In contrast, increasing
       concentrations of Na ions up to 5 M were without effect. Increasing pH
       had similar effects on HIV-1 protease as increasing Mg2+ ions
       concentration, with concomitant dissociation into subunits, increase in
       quantum yield and red shift in tryptophan fluorescence, and loss in
       catalytic activity. The apparent pKa for these structural and functional
       transitions was 6.95 +/- 0.08. This value is consistent with that of an
       aspartic acid residue with an anomalously high pKa, which has been
       implicated in the catalytic activity of HIV-1 protease.
 DE    Amino Acid Sequence  Cations, Divalent  Chromatography, Gel  Comparative
       Study  Escherichia coli  Hydrogen-Ion Concentration  Hydrolysis  HIV
       Protease/*CHEMISTRY/METABOLISM  Macromolecular Systems
       Magnesium/METABOLISM  Magnesium Chloride/*PHARMACOLOGY  Molecular
       Sequence Data  Recombinant Proteins/CHEMISTRY  Sodium
       Chloride/PHARMACOLOGY  Support, Non-U.S. Gov't  Support, U.S. Gov't,
       P.H.S.  JOURNAL ARTICLE

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