An Obligate Domain-Swapped Dimer of Cyanovirin with Enhanced Anti-Viral Activity
Description of Invention:
The present invention provides a purified or isolated obligate domain-swapped dimer of Cyanovirin-N (CVN hereafter), a method of making an obligate domain-swapped dimer of CVN and a method of inhibiting a viral infection of a mammal by administering domain-swapped dimer of CVN. CVN is outstanding in that it potently blocks viral entry in all human and simian isolates by binding to HIV through highly avid and very specific carbohydrate-mediated interactions with the surface envelope glycoprotein gp120. CVN has also been shown to form a domain-swapped dimer under non-physiological conditions such as mM concentration and low pH. This invention provides an obligate domain-swapped dimeric mutant of CVN, called deltaQ50-CVN, which has several significant advantages over the wild-type CVN: First, deltaQ50-CVN can be purified form a crude bacterial cell lysate in a single chromatographic step because it forms only one homogeneous species. Second, because this obligate dimer has four carbohydrate binding sites, it binds gp120 and other glycoproteins with greater affinity than wild-type CVN. Third, deltaQ50-CVN shows an enhancing increase in efficacy in blocking viral entry in a quantitative HIV-1 envelope-mediated cell fusion assay. Thus, deltaQ50-CVN displays enhanced anti-HIV activity relative to the wild-type CVN monomer and offers a great advantage over wild-type CVN because it is extremely easy to purify large quantities to greater than 95% homogeneity. So, it may open the possibility that an effective drug treatment for HIV could reach underdeveloped countries.
Inventors:
Carole A. Bewley and Brendans Kelly (NIDDK)
Patent Status:
DHHS Reference No. E-096-02/0 filed 25 Feb 2002
Relevant Publication:
Bewley, C.A. (2001) Solution structure of a cyanovirin-N:Manalpha1-2Manalpha complex: structural basis for high affinity carbohydrate-mediated binding to the HIV-envelope protein gp120. Structure, 10, 931-940.
Clore, G.M. & Bewley, C.A. (2002) Using conjoined rigid body / torsion angle simulated annealing to determine the relative orientation of covalently linked protein domains from dipolar couplings. J. Magn. Reson. 154: 329-335.
Bewley, C.A. and Otero-Quintero, S. (2001) The potent anti-HIV protein cyanovirin-N contains two novel carbohydrate binding sites that selectively bind to Man8 D1D3 and Man9 with nanomolar affinity: implications for binding to the HIV envelope glycoprotein gp120. J. Am. Chem. Soc. 123: 3892-3902.
Bewley, C.A. and Clore, G.M. (2000) Determination of the relative orientation of the two halves of the domain-swapped dimer of cyanovirin-N in solution using dipolar couplings and rigid body minimzation. J. Am. Chem. Soc. 122: 6009-6016.
Yang, F., Bewley, C.A., Louis, J.M., Gustafson, K.R., Boyd, M.R., Gronenborn, A.M., Clore, G.M., Wlodawer, A. (1999) Crystal structure of cyanovirin-N, a potent HIV-inactivating protein shows unexpected domain-swapping. J. Mol. Biol. 288: 403-412.
For Additional Information Please Contact: Sally Hu PhD MBA
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