Mori,T.; Barrientos,L.G.; Han,Z.Z.; Gronenborn,A.M.; Turpin,J.A.; Boyd,M.R.: Protein Expr.Purif. 26: 42-49, 2002.
Abstract:
Cyanovirin-N (CV-N) is under development as a topical (vaginal or rectal) microbicide to prevent sexual transmission of human immunodeficiency virus (HIV), and an economically feasible means for very large-scale production of the protein is an urgent priority. We observed that N-glycosylation of CV-N in yeast eliminated the anti-HIV activity, and that dimeric forms or aggregates of CV-N occurred under certain conditions, potentially complicating the efficient, large-scale manufacture of pure monomeric CV-N. Therefore, we have expressed and compared CV-N homologs in which the glycosylation-susceptible Asn residue at position 30 was replaced with Ala,Gln or Val, and/or the Pro at position 51 was replaced by Gly to eliminate a potential site of cis-trans isomerization. All of the homologs had anti-HIV activity comparable to wild-type CV-N, and the Pro51Gly homologs exhibited enhanced stability toward denaturants. These glycosylation-resistant,functional cyanovirins are amenable to large-scale production either in bacteria or in eukaryotic hosts such as yeasts or transgenic plants or animals.
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