Sortase A as a tool for high‐yield histatin cyclization

• Published in: The FASEB journal Vol. 25; no. 8; pp. 2650 - 2658
• Main Authors: Bolscher, Jan G. M., Oudhoff, Menno J., Nazmi, Kamran, Antos, John M., Guimaraes, Carla P., Spooner, Eric, Haney, Evan F., Vallejo, Juan J. Garcia, Vogel, Hans J., Hof, Wim van't, Ploegh, Hidde L., Veerman, Enno C. I.
• Format: Journal Article
• Language: English
• Published: United States 01-08-2011
• Subjects: Amino Acid Sequence; Aminoacyltransferases - genetics; Aminoacyltransferases - metabolism; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Cell Line; Circular Dichroism; Cysteine Endopeptidases - genetics; Cysteine Endopeptidases - metabolism; Histatins - biosynthesis; Histatins - chemistry; Histatins - genetics; Histatins - pharmacology; Humans; intramolecular transpeptidation; Molecular Sequence Data; Nuclear Magnetic Resonance, Biomolecular; Peptides, Cyclic - biosynthesis; Peptides, Cyclic - chemistry; Peptides, Cyclic - genetics; Peptides, Cyclic - pharmacology; Protein Conformation; Protein Engineering; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Recombinant Proteins - pharmacology; Staphylococcus aureus - enzymology; Staphylococcus aureus - genetics; wound closure stimulating peptide; Wound Healing - drug effects
• ISSN: 0892-6638; 1530-6860
• DOI: 10.1096/fj.11-182212

ABSTRACT
Cyclic peptides are highly valued tools in biomedical research. In many cases, they show higher receptor affinity, enhanced biological activity, and improved serum stability. Technical difficulties in producing cyclic peptides, especially larger ones, in appreciable yields have precluded a prolific use in biomedical research. Here, we describe a novel and efficient cyclization method that uses the peptidyl‐transferase activity of the Staphylococcus aureus enzyme sortase A to cyclize linear synthetic precursor peptides. As a model, we used histatin 1, a 38‐mer salivary peptide with motogenic activity. Chemical cyclization of histatin 1 resulted in ≤3% yields, whereas sortase‐mediated cyclization provided a yield of >90%. The sortase‐cyclized peptide displayed a maximum wound closure activity at 10 nM, whereas the linear peptide displayed maximal activity at 10 μM. Circular dichroism and NMR spectroscopic analysis of the linear and cyclic peptide in solution showed no evidence for conformational changes, suggesting that structural differences due to cyclization only became manifest when these peptides were located in the binding domain of the receptor. The sortase‐based cyclization technology provides a general method for easy and efficient manufacturing of large cyclic peptides.—Bolscher, J. G. M., Oudhoff, M. J., Nazmi, K., Antos, J. M., Guimaraes, C. P., Spooner, E., Haney, E. F., Garcia‐Vallejo, J. J., Vogel, H. J., van't Hof, W., Ploegh, H. L., Veerman. E. C. I. Sortase A as a tool for high‐yield histatin cyclization. FASEB J. 25, 2650–2658 (2011). www.fasebj.org