Shenoy,S.R.; O'Keefe,B.R.; Bolmstedt,A.J.; Cartner,L.K.; Boyd,M.R.: J.Pharmacol.Exp.Ther. 297: 704-710, 2001.
Abstract:
The virucidal protein cyanovirin-N (CV-N) mediates its highly potent anti-HIV activity, at least in part, through interactions with the viral envelope glycoprotein, gp120. Here we dissect in further detail the mechanism of CV-N's glycosylation-dependent binding to gp120. Isothermal titration calorimetry (ITC) binding studies of CV-N with Endo H-treated gp120 showed that binding was completely abrogated by removal of high-mannose oligosaccharides from the glycoprotein. Additional ITC and CDS spectral studies with CVC-N and other glycoproteins as well showed that CV-N discriminately bound only glycoproteins that contain high-mannose oligosaccharides. Binding experiments with RNase B indicated that the single high-mannose oligosaccharide on that enzyme mediated all of its binding with CV-N (Kd 0.602 µM). A finer level of oligosaccharide selectivity of CV-N was revealed in affinity chromatography-LCMS experiments which showed that CV-N preferentially bound only oligomannose-8 (Man-8) and oligomannose-9 (Man-9) isoforms of RNase B. Finally we biophysically characterized the interaction of CV-N with purified, single oligosaccharide, Man-8. The binding affinity of Man-8 for CV-N is unusually strong (Kd 0.488 µM), several hundred-fold greater than observed for oligosaccharides and their protein lectins (Kd 1 mM-1µM), further establishing a critical role of high-mannose oligosaccharides in CV-N binding to glycoproteins.
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