High throughput screening for cyanovirin-N mimetics binding to HIV-1 gp41.
The human immunodeficiency virus type-1 (HIV-1) envelope glycoprotein gp41 is an important mediator of viral entry into host cells. Previous studies showed that the virucidal protein cyanovirin-N (CV-N) bound to both gp120 and gp41, and that this binding was associated with its antiviral activity. We constructed an HTS assay based on the interaction of europium-labeled CV-N with recombinant glycosylated gp41 ectodomain to support identification of small-molecule mimetics of CV-N that might be developed as antiviral drug leads. Primary screening of over 107,000 natural product extracts in the assay yielded 347 confirmed hits. Secondary assays eliminated extracts that bound directly to labeled CV-N or for which the simple sugars mannose and N-acetylglucosamine blocked the interaction with gp41 (lectin activity). Extracts were further prioritized based on anti-HIV activity and other biological, biochemical, and chemical criteria. The distribution of source organism taxonomy of active extracts was analyzed, as was the cross-correlation of activity between the CV-N-gp41 binding competition assay and the previously reported CV-N-gp120 binding competition assay. A limited set of extracts was selected for bioassay-guided fractionation.
Beutler
John A
JA
Intramural Research Support Program and Molecular Targets Drug Discovery Program, SAIC-Frederick, Center for Cancer Research, National Cancer Institute-Frederick, Frederick, MD 21702-1201, USA.
McMahon
James B
JB
Johnson
Tanya R
TR
O'Keefe
Barry R
BR
Buzzell
Randy A
RA
Robbins
Danielle
D
Gardella
Roberta
R
Wilson
Jennifer
J
Boyd
Michael R
MR
D000117:Acetylglucosamine / Q000378:metabolism; D019380:Anti-HIV Agents / Q000494:pharmacology*; D001426:Bacterial Proteins; D001667:Binding, Competitive; D001681:Biological Assay; D001685:Biological Factors; D002352:Carrier Proteins / Q000378:metabolism / Q000494:pharmacology*; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004797:Enzyme-Linked Immunosorbent Assay; D015699:HIV Envelope Protein gp120 / Q000378:metabolism; D015700:HIV Envelope Protein gp41 / Q000737:chemistry / Q000378:metabolism*; D015497:HIV-1 / Q000378:metabolism; D006801:Humans; D008358:Mannose / Q000378:metabolism; D009005:Monosaccharides / Q000378:metabolism; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods
D019380:Anti-HIV Agents; D001426:Bacterial Proteins; D001685:Biological Factors; D002352:Carrier Proteins; D015699:HIV Envelope Protein gp120; D015700:HIV Envelope Protein gp41; D009005:Monosaccharides; D011994:Recombinant Proteins; C107733:cyanovirin N; D008358:Mannose; D000117:Acetylglucosamine