Lysine-scanning mutagenesis reveals an amendable face of the cyclotide kalata B1 for the optimization of nematocidal activity

Huang, Yen-Hua (黃彥華), Colgrave, Michelle L., Clark, Richard J., Kotze, Andrew C. and Craik, David J. (2010) Lysine-scanning mutagenesis reveals an amendable face of the cyclotide kalata B1 for the optimization of nematocidal activity. Journal of Biological Chemistry, 285 14: 10797-10805. doi:10.1074/jbc.M109.089854

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Author Huang, Yen-Hua (黃彥華)
Colgrave, Michelle L.
Clark, Richard J.
Kotze, Andrew C.
Craik, David J.
Title Lysine-scanning mutagenesis reveals an amendable face of the cyclotide kalata B1 for the optimization of nematocidal activity
Journal name Journal of Biological Chemistry   Check publisher's open access policy
ISSN 0021-9258
1083-351X
Publication date 2010-04-02
Sub-type Article (original research)
DOI 10.1074/jbc.M109.089854
Open Access Status File (Publisher version)
Volume 285
Issue 14
Start page 10797
End page 10805
Total pages 9
Editor Herbert Tabor
Place of publication Bethesda, MD, U.S.A.
Publisher American Society for Biochemistry and Molecular Biology
Collection year 2011
Language eng
Formatted abstract
Cyclotides are a family of macrocyclic peptides that combine the unique features of a head-to-tail cyclic backbone and a cystine knot motif, the combination of which imparts them with extraordinary stability. The prototypic cyclotide kalata B1 is toxic against two economically important gastrointestinal nematode parasites of sheep, Haemonchus contortus and Trichostrongylus colubriformis. A lysine scan was conducted to examine the effect of the incorporation of positive charges into the kalata B1 cyclotide framework. Each of the non-cysteine residues in this 29-amino acid peptide was successively substituted with lysine, and the nematocidal and hemolytic activities of the suite of mutants were determined. Substitution of 11 residues within kalata B1 decreased the nematocidal activity dramatically. On the other hand, six other residues that are clustered on the surface of kalata B1 were tolerant to Lys substitution, and indeed the introduction of positively charged residues into this region increased nematocidal activity. This activity was increased further in double and triple lysine mutants, with a maximal increase (relative to the native kalata B1) of 13-fold obtained with a triple lysine mutant (mutated at positions Thr-20, Asn-29, and Gly-1). Hemolytic activity correlated with the nematocidal activity of all lysine mutants. Our data clearly highlight the residues crucial for nematocidal and hemolytic activity in cyclotides, and demonstrate that the nematocidal activity of cyclotides can be increased by incorporation of basic amino acids.
© 2010 by The American Society for Biochemistry and Molecular Biology, Inc.
Keyword Cyclic-cystine-knot
Virus 3C protease
Plant cyclotides
Momordica-cochinchinensis
Insecticidal activity
Haemonchus-contortus
Macrocyclic peptides
Biological-activity
Structural motif
Cycloviolacin O2
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2011 Collection
Institute for Molecular Bioscience - Publications
 
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Created: Sun, 18 Apr 2010, 00:04:14 EST