Skip NavigationSkip to Content
Return Return to Search Results

Topology of the disulfide bonds in the antiviral lectin scytovirin

  1. Author:
    Moulaei, T.
    Stuchlik, O.
    Reed, M.
    Yuan, W. R.
    Pohl, J.
    Lu, W. Y.
    Haugh-Krumpe, L.
    O'Keefe, B. R.
    Wlodawer, A.

  2. Author Address

    [Moulaei, Tinoush; Wlodawer, Alexander] NCI, Prot Struct Sect, Macromol Crystallog Lab, Frederick, MD 21702 USA. [Stuchlik, Olga; Reed, Matthew; Pohl, Jan] Ctr Dis Control & Prevent, Biotechnol Core Facil Branch, Div Sci Resources, Atlanta, GA 30333 USA. [Yuan, Weirong; Lu, Wuyuan] Univ Maryland, Sch Med, Inst Human Virol, Dept Biochem & Mol Biol, Baltimore, MD 21201 USA. [Haugh-Krumpe, Lauren] NCI, SAIC Frederick Inc, Frederick, MD 21702 USA. [O'Keefe, Barry R.] NCI, Mol Targets Lab, Ctr Canc Res, Frederick, MD 21702 USA.;Wlodawer, A, NCI, Prot Struct Sect, Macromol Crystallog Lab, Frederick, MD 21702 USA.;wlodawer@nih.gov
    1. Year: 2010
    2. Date: Sep
  2. Journal: Protein Science
    1. 19
    2. 9
    3. Pages: 1649-1661
  4. Type of Article: Article
  5. ISSN: 0961-8368
  1. Abstract:

    The antiviral lectin scytovirin (SVN) contains a total of five disulfide bonds in two structurally similar domains. Previous reports provided contradictory results on the disulfide pairing in each individual domain, and we have now re-examined the disulfide topology. N-terminal sequencing and mass spectrometry were used to analyze proteolytic fragments of native SVN obtained at acidic pH, yielding the assignment as Cys7-Cys55, Cys20-Cys32, Cys26-Cys38, Cys68-Cys80, and Cys74-Cys86. We also analyzed the N-terminal domain of SVN (SD1, residues 1-48) prepared by expression/oxidative folding of the recombinant protein and by chemical synthesis. The disulfide pairing in the chemically synthesized SD1 was forced into predetermined topologies: SD1A (Cys20-Cys26, Cys32-Cys38) or SD1 B (Cys20-Cys32, Cys26-Cys38). The topology of native SVN was found to be in agreement with the SD1B and the one determined for the recombinant SD1 domain. Although the two synthetic forms of SD1 were distinct when subjected to chromatography, their antiviral properties were indistinguishable, having low nM activity against HIV. Tryptic fragments, the "cystine clusters" [Cys20-Cys32/Cys26-Cys38; SD1] and [Cys68-Cys80/Cys74-C-86; SD2], were found to undergo rapid disulfide interchange at pH 8. This interchange resulted in accumulation of artifactual fragments in alkaline pH digests that are structurally unrelated to the original topology, providing a rational explanation for the differences between the topology reported herein and the one reported earlier (Bokesh et al., Biochemistry 2003;42:2578-2584). Our observations emphasize the fact that proteins such as SVN, with disulfide bonds in close proximity, require considerable precautions when being fragmented for the purpose of disulfide assignment.

    See More

  1. Keywords:

External Sources

  1. View Full Text
  2. DOI: 10.1002/pro.445
  3. View record in Web of Science®
  4. WOS: 000281563000005

Library Notes

  1. Fiscal Year: FY2009-2010
NCI at Frederick

You are leaving a government website.

This external link provides additional information that is consistent with the intended purpose of this site. The government cannot attest to the accuracy of a non-federal site.

Linking to a non-federal site does not constitute an endorsement by this institution or any of its employees of the sponsors or the information and products presented on the site. You will be subject to the destination site's privacy policy when you follow the link.

ContinueCancel