Essential regions for antiviral activities of actinohivin, a sugar-binding anti-human immunodeficiency virus protein from an actinomycete.
Actinohivin (AH) is a potent anti-human immunodeficiency virus (HIV) protein that consists of highly conserved three-tandem repeats (segments 1, 2, and 3). The molecular target of AH in its anti-HIV activity is high-mannose-type saccharide chains of HIV gp120. This article deals with sequence requirements for the anti-HIV activity of AH. The deleted or substituted DNAs encoding AH or His-AH were prepared using mutagenic oligonucleotide primers in PCR. The mutant constructs were expressed in Escherichia coli, and the activities of the recombinant protein products were examined by a syncytium-formation assay system that mimics anti-HIV activity. The single segment mutant His-AHs showed no anti-syncytium-formation activity, but the mutant His-AHs, which consists of 2 or 3 segments, retained reduced activities. His-AH(6-114) dramatically reduced the anti-syncytium-formation activity to that of His-AH(36-114) or His-AH(I5A). Furthermore, His-AH(Q33A), His-AH(Q71A), and His-AH(Q109A) in which glutamine residues were substituted into alanine showed reduced activities of 1/20, 1/10, and 1/30, respectively, in anti-syncytium formation compared with His-AH. These results indicate that three segments of AH are necessary for potent anti-syncytium-formation activity-that is, for potent anti-HIV activity and the cooperated involvement of each segment of AH increased the AH-gp120 interaction.
Takahashi
Atsushi
A
School of Pharmaceutical Sciences, Kitasato University, 5-9-1 Shirokane, Minato-ku, Tokyo 108-8641, Japan.
Inokoshi
Junji
J
Chiba
Harumi
H
Omura
Satoshi
S
Tanaka
Haruo
H
D039903:Actinobacteria / Q000737:chemistry*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D019380:Anti-HIV Agents / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D050260:Carbohydrate Metabolism; D017124:Conserved Sequence; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D015726:Giant Cells / Q000187:drug effects; D006678:HIV / Q000187:drug effects* / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D020128:Inhibitory Concentration 50; D008969:Molecular Sequence Data; D017384:Sequence Deletion / Q000235:genetics
D019380:Anti-HIV Agents; D001426:Bacterial Proteins; C424136:actinohivin protein, actinomycete; D003545:Cysteine