Visualizing the molecular interactions of a nucleotide analog, GS-9148, with HIV-1 reverse transcriptase-DNA complex.

Abstract:

GS-9148 ([5-(6-amino-purin-9-yl)-4-fluoro-2,5-dihydro-furan-2-yloxymethyl]-phospho nic acid) is a dAMP (2'-deoxyadenosine monophosphate) analog that maintains its antiviral activity against drug-resistant HIV. Crystal structures for HIV-1 reverse transcriptase (RT) bound to double-stranded DNA, ternary complexes with either GS-9148-diphosphate or 2'-deoxyadenosine triphosphate (dATP), and a post-incorporation structure with GS-9148 translocated to the priming site were obtained to gain insight into the mechanism of RT inhibition. The binding of either GS-9148-diphosphate or dATP to the binary RT-DNA complex resulted in the fingers subdomain closing around the incoming substrate. This produced up to a 9 A shift in the tips of the fingers subdomain as it closed toward the palm and thumb subdomains. GS-9148-diphosphate shows a similar binding mode as dATP in the nucleotide-binding site. Residues whose mutations confer resistance to nucleotide/nucleoside RT inhibitors, such as M184, Y115, L74, and K65, show little to no shift in orientation whether GS-9148-diphosphate or dATP is bound. One difference observed in binding is the position of the central ring. The dihydrofuran ring of GS-9148-diphosphate interacts with the aromatic side chain of Y115 more than does the ribose ring of dATP, possibly picking up a favorable pi-pi interaction. The ability of GS-9148-diphosphate to mimic the active-site contacts of dATP may explain its effective inhibition of RT and maintained activity against resistance mutations. Interestingly, the 2'-fluoro moiety of GS-9148-diphosphate was found in close proximity to the Q151 side chain, potentially explaining the observed moderately reduced susceptibly to GS-9148 conferred by Q151M mutation.

Polymerases:

Topics:

Nucleotide Analogs / Template Lesions, Structure and Structure/Function, Nucleotide Incorporation, Reverse Transcriptase, Source / Purification

Status:

new topics/pols set partial results complete validated

Results:

Polymerase Reference Property Result Context
HIV RT Lansdon EB2010 Reverse Transcriptase Activity Yes
HIV RT Lansdon EB2010 Polymerase Catalytic Residue Amino Acids D110, D185, D186
HIV RT Lansdon EB2010 Incorporation of non-standard nucleotides Unspecified
HIV RT Lansdon EB2010 Cloned or native Cloned in E. coli
HIV RT Lansdon EB2010 Tagged Yes
HIV RT Lansdon EB2010 Tag Name his
HIV RT Lansdon EB2010 Residues Involved in Catalysis of RNase H D443, E478, D498, D549
HIV RT Lansdon EB2010 Full length or truncated Full length

Structures:

3KK3 3KK2 3KK1 3KJV
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