Abstract
It remains difficult to obtain high-resolution atomic force microscopy images of HIV-1 integrase bound to DNA in a dimeric or tetrameric fashion. We therefore constructed specific target DNAs to assess HIV-1 integrase binding and purified the complex by dialysis prior to analysis. Our resulting atomic force microscopy analyses indicated precise size of binding human immunodeficiency virus type 1 (HIV-1) recombinant integrase in a tetrameric manner, inducing formation of a loop-like or figure-eight-like secondary structure in the target DNA. Our findings regarding the target DNA secondary structure provide new insights into the intermediate states of retroviral integration.
Highlights
Retroviruses integrate their genome into the genome of host cells in a process catalyzed by the enzyme integrase, which binds the 59- and 39- termini of retroviral long terminal repeats (LTRs) and integrates them into the host cell’s DNA
Our study revealed that oligomeric integrase digests the target DNA at the oligomeric integrase-binding site with secondary structure formation
atomic force microscopy (AFM) analysis indicated that human immunodeficiency virus type 1 (HIV-1) recombinant integrase bound to the target DNA xkyk
Summary
Retroviruses integrate their genome into the genome of host cells in a process catalyzed by the enzyme integrase, which binds the 59- and 39- termini of retroviral long terminal repeats (LTRs) and integrates them into the host cell’s DNA. Elucidating the underlying mechanism of integration will necessitate that we first obtain a more complete understanding of the transitional states involved in the binding of HIV-1 integrase to target DNA. Hare et al recently reported a crystal structure of prototype foamy virus integrase complexed with its cognate DNA [1] that is suggestive of a tetrameric structure. HIV-1 integrase forms stable tetramers and associates with the transcriptional coactivator LEDGF/p75, which is an essential cofactor for HIV integration [2,3]. Inhibitors have been developed to target tetrameric integrases [4,5]. No convincing visualizations of integrase tetramerization have been reported, perhaps due to difficulties in generating a tetramer stable enough to visualize
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