Fine Structure Target Analysis Using

The target specificity of DNA strand transfer mediated by human immunodeficiency virus type 1 integrase was examined in vitro with synthetic oligonucleotides. Although insertion occurred at most locations in the target, some sites were preferred over others by at least 15-fold. Changing the nucleotide sequence of the target changed the distribution of preferred sites in complex ways, some of which included changes in target preference distant from the sequence alteration. Alignment of target sequences revealed that adenosine is preferred adjacent to the insertion site. Strand transfer occurred to within 2 nucleotides of the 3 ' end and to within 3 nucleotides of the 5 ' end of the target. This suggests that only 2 or 3 nucleotides flanking the target site are required for integration; such restricted contact with target DNA would allow integrase to insert the two ends of viral DNA into two closely spaced sites in host DNA, consistent with the concerted in vivo integration reaction that generates a 5-bp target duplication. Insertion of viral DNA into host DNA is an integral part of retroviral life cycles and occurs at many chromosomal locations. Integration is mediated by the viral integrase, which removes (trims) 2 nucleotides from the 3 ' end of each linear viral DNA strand and transfers these recessed 3 ' end