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Microwave-assisted synthesis of triple-helical, collagen-mimetic lipopeptides

Nature Protocols volume 5, pages 39–50 (2010)Cite this article

Abstract

Collagen-mimetic peptides and lipopeptides are widely used as substrates for matrix degrading enzymes, as new biomaterials for tissue engineering, as drug delivery systems and so on. However, the preparation and subsequent purification of these peptides and their fatty-acid conjugates are really challenging. Herein, we report a rapid microwave-assisted, solid-phase synthetic protocol to prepare the fatty-acid conjugated, triple-helical peptides containing the cleavage site for the enzyme matrix metalloproteinase-9 (MMP-9). We employed a PEG-based resin as the solid support and the amino acids were protected with Fmoc- and tert-butyl groups. The amino acids were coupled at 50 °C (25 W of microwave power) for 5 min. The deprotection reactions were carried out at 75 °C (35 W of microwave power) for 3 min. Using this protocol, a peptide containing 23 amino acids was synthesized and then conjugated to stearic acid in 14 h.

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Figure 9: Circular dichroism (CD) spectrum of the purified lipopepide recorder at 25 °C.
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Acknowledgements

This research was supported by the NIH grant 1R01 CA113746, and NSF DMR-0705767 to S.M. J.B. is supported by a Graduate School Doctoral Fellowship from the North Dakota State University. J.B.S. thanks the support of the proteomics core facility by NIH Grant P20 RR016741 from the INBRE Program of the NCRR.

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Authors and Affiliations

  1. Department of Pharmaceutical Sciences, North Dakota State University, Fargo, North Dakota, USA

    Jayati Banerjee, Andrea J Hanson & Sanku Mallik

  2. Department of Biochemistry and Molecular Biology, School of Medicine and Health Sciences, University of North Dakota, Grand Forks, North Dakota, USA

    Wallace W Muhonen & John B Shabb

Authors
  1. Jayati Banerjee
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  2. Andrea J Hanson
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  3. Wallace W Muhonen
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Contributions

S.M. conceived the idea and supervised the project; J.B. and A.J.H. carried out the lipopeptide synthesis; J.B.S. and W.W.M. recorded the mass spectra; and S.M. and J.B. wrote the manuscript.

Corresponding author

Correspondence to Sanku Mallik.

Supplementary information

Supplementary Methods

Detailed operational steps the CEM Liberty peptide synthesizer performs for the single coupling, double coupling and cleavage from the resin. (PDF 75 kb)

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Banerjee, J., Hanson, A., Muhonen, W. et al. Microwave-assisted synthesis of triple-helical, collagen-mimetic lipopeptides. Nat Protoc 5, 39–50 (2010). https://doi.org/10.1038/nprot.2009.195

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