Multivalent interactions with gp120 are required for the anti‐HIV activity of Cyanovirin

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Abstract
Cyanovirin‐N (CV‐N) is a cyanobacterial lectin that binds to specific oligomannoses on the surface of gp120, resulting in nanomolar antiviral activity against HIV. In its monomeric form, CV‐N contains two functional carbohydrate‐binding domains, A and B. When refolded at high concentration, the protein can form a domain‐swapped dimer. To clarify the role of multiple‐binding sites in CV‐N, we previously designed a monomeric mutant, P51G‐m4‐CVN, in which the binding site on domain A was rendered ineffective by four mutations (m4); in addition, a hinge region mutation (P51G) hinders the formation of a domain swapped dimer. The protein bound gp120 with diminished affinity and was completely inactive against HIV. Here, we present two mutants, ΔQ50‐m4‐CVN and S52P‐m4‐CVN, which fold exclusively as domain‐swapped dimers while containing the four mutations that abolish domain A. The dimers contain two intact B domains, thus restoring multivalency. ΔQ50‐m4‐CVN and S52P‐m4‐CVN bind gp120 at low‐nanomolar concentrations and recover in part the antiviral activity of wt CV‐N. These results indicate that the number of carbohydrate binding domains, rather than their identity, is crucial to CV‐N functionality. © 2009 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 92: 194–200, 2009.This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com