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Journal of Virology, August 2001, p. 7756-7762, Vol. 75, No. 16
Department of Microbiology and
Immunology1 and Department of
Medicine,2 Pennsylvania State University
College of Medicine, The Milton S. Hershey Medical Center, Hershey,
Pennsylvania 17033
Received 13 December 2000/Accepted 21 May 2001
To identify parts of retroviral integrase that interact with
cellular DNA, we tested patient-derived human immunodeficiency virus
type 1 (HIV-1) integrases for alterations in the choice of nonviral
target DNA sites. This strategy took advantage of the genetic diversity
of HIV-1, which provided 75 integrase variants that differed by a small
number of amino acids. Moreover, our hypothesis that biological
pressures on the choice of nonviral sites would be minimal was
validated when most of the proteins that catalyzed DNA joining
exhibited altered target site preferences. Comparison of the sequences
of proteins with the same preferences then guided mutagenesis of a
laboratory integrase. The results showed that single amino acid
substitutions at one particular residue yielded the same target site
patterns as naturally occurring integrases that included these
substitutions. Similar results were found with DNA joining reactions
conducted with Mn2+ or with Mg2+ and were
confirmed with a nonspecific alcoholysis assay. Other amino acid
changes at this position also affected target site preferences. Thus,
this novel approach has identified a residue in the central domain of
HIV-1 integrase that interacts with or influences interactions with
cellular DNA. The data also support a model in which integrase has
distinct sites for viral and cellular DNA.
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.16.7756-7762.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Use of Patient-Derived Human Immunodeficiency Virus Type 1 Integrases To Identify a Protein Residue That Affects Target
Site Selection
*
Corresponding author. Mailing address: Department of
Medicine, Division of Infectious Diseases, Pennsylvania State
University College of Medicine, The Milton S. Hershey Medical Center,
P.O. Box 850, Mail Services H036, Hershey, PA 17033-0850. Phone: (717) 531-8881. Fax: (717) 531-4633. E-mail: mkatzman{at}psu.edu.
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