A new "Era" for cyclotide sequencing

Colgrave, Michelle L., Poth, Aaron G., Kaas, Quentin and Craik, David J. (2010). A new "Era" for cyclotide sequencing. In: 1st International Conference on Circular Proteins. Procceedings.. 1st International Conference on Circular Proteins, Heron Island, Qld, Australia, (592-601). 18-21 October 2009. doi:10.1002/bip.21400


Author Colgrave, Michelle L.
Poth, Aaron G.
Kaas, Quentin
Craik, David J.
Title of paper A new "Era" for cyclotide sequencing
Conference name 1st International Conference on Circular Proteins
Conference location Heron Island, Qld, Australia
Conference dates 18-21 October 2009
Proceedings title 1st International Conference on Circular Proteins. Procceedings.   Check publisher's open access policy
Journal name Biopolymers - Peptide Science   Check publisher's open access policy
Place of Publication New York, NY, United States
Publisher John Wiley & Sons
Publication Year 2010
Year available 2010
Sub-type Fully published paper
DOI 10.1002/bip.21400
ISSN 0006-3525
1097-0282
Volume 94
Issue 5
Start page 592
End page 601
Total pages 10
Collection year 2011
Language eng
Abstract/Summary In recent years, the discovery of a large family of macro cyclic peptides, the cyclotides, has revealed Nature's ingenuity in molecular drug design. The incorporation of a cyclic peptide backbone and a knotted arrangement of disulfide bridges into their structures confers extraordinary chemical, thermal, and enzymatic stability on these biologically active peptides. However, these structural attributes present challenges in the identification of cyclotides. Until now, the sequencing of cyclotides has been slow and inefficient owing to inherent difficulties in the separation of these hydrophobic peptides from plants, the multiple chemical and enzymatic derivatization steps required to make them amenable to mass spectrometric sequencing, and the lack of software tools to efficiently deal with these circular permutants. The current bottleneck slowing the speed of cyclotide sequencing is the requirement for multiple HPLC purification steps before analysis. Here, we have applied proteomic strategies to fast-track the discovery of known, modified and novel sequences. Using four fractions from a previously well-characterized cyclotide-containing plant species, Viola odorata, 11 new sequences, as well as a plethora of known and modified cyclotides, were uncovered. In addition, the methodology was validated through analysis of crude leaf extracts of Oldenlandia affinis and Arabidopsis thaliana. The unambiguous identification of a suite of cyclotides in the Oldenlandia affinis extract provided the ultimate proof-of-concept for this application. Major advances in methodology include the use of optimized LC-MS/MS conditions and design of a custom-built cyclotide database, in which mature cyclotide sequences are excised, replicated and appended, marking a new "era" for cyclotide sequencing. 2010 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 94: 592-601, 2010.
Keyword Cyclotide
Sequencing
Mass spectrometry
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ
Additional Notes Special Issue: Special Issue on International Conference on Circular Proteins, Article first published online: 4 August, 2010.

 
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Created: Tue, 30 Nov 2010, 13:47:13 EST by Dr Quentin Kaas on behalf of Institute for Molecular Bioscience