Document 0128
 DOCN  M94A0128
 TI    Folding of the multidomain human immunodeficiency virus type-I
       integrase.
 DT    9412
 AU    Grandgenett DP; Goodarzi G; Institute for Molecular Virology, St. Louis
       University, Missouri; 63110.
 SO    Protein Sci. 1994 Jun;3(6):888-97. Unique Identifier : AIDSLINE
       MED/94348418
 AB    Protein folding conditions were established for human immunodeficiency
       virus integrase (IN) obtained from purified bacterial inclusion bodies.
       IN was denatured by 6 M guanidine.HCl-5 mM dithiothreitol, purified by
       gel filtration, and precipitated by ammonium sulfate. The reversible
       solvation of precipitated IN by 6 M guanidine.HCl allowed for wide
       variation of protein concentration in the folding reaction. A 6-fold
       dilution of denatured IN by 1 M NaCl buffer followed by dialysis
       produced enzymatically active IN capable of 3' OH end processing, strand
       transfer, and disintegration using various human immunodeficiency
       virus-1 (HIV-1) long terminal repeat DNA substrates. The specific
       activities of folded IN preparations for these enzymatic reactions were
       comparable to those of soluble IN purified directly from bacteria. The
       subunit composition and enzymatic activities of IN were affected by the
       folding conditions. Standard folding conditions were defined in which
       monomers and protein aggregates sedimenting as dimers and tetramers wree
       produced. These protein aggregates were enzymatically active, whereas
       monomers had reduced strand transfer activity. Temperature modifications
       of the folding conditions permitted formation of mainly monomers. Upon
       assaying, these monomers were efficient for strand transfer and
       disintegration, but the oligomeric state of IN under the conditions of
       the assay is determinate. Our results suggest that monomers of the
       multidomain HIV-1 IN are folded correctly for various catalytic
       activities, but the conditions for specific oligomerization in the
       absence of catalytic activity are undefined.
 DE    Ammonium Sulfate  Blotting, Western  Chromatography, Gel  Dithiothreitol
       DNA Nucleotidyltransferases/*CHEMISTRY/METABOLISM  DNA, Viral/METABOLISM
       Enzyme Stability  Freezing  Guanidines  Heat  HIV Long Terminal Repeat
       HIV-1/*ENZYMOLOGY  Precipitation  Protein Denaturation  *Protein Folding
       Recombinant Proteins/CHEMISTRY/METABOLISM  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).