title: Sortase A as a tool for high-yield histatin cyclization.
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.
authors.affiliation: Department of Oral Biochemistry, Academic Centre for Dentistry Amsterdam, University of Amsterdam and VU Amsterdam, Gustav Mahlerlaan 3004, 1081 LA, Amsterdam, The Netherlands. j.bolscher@acta.nl
authors.forename: Jan G M
authors.initials: JG
authors.lastname: Bolscher
authors.forename: Menno J
authors.initials: MJ
authors.lastname: Oudhoff
authors.forename: Kamran
authors.initials: K
authors.lastname: Nazmi
authors.forename: John M
authors.initials: JM
authors.lastname: Antos
authors.forename: Carla P
authors.initials: CP
authors.lastname: Guimaraes
authors.forename: Eric
authors.initials: E
authors.lastname: Spooner
authors.forename: Evan F
authors.initials: EF
authors.lastname: Haney
authors.forename: Juan J
authors.initials: JJ
authors.lastname: Garcia Vallejo
authors.forename: Hans J
authors.initials: HJ
authors.lastname: Vogel
authors.forename: Wim
authors.initials: W
authors.lastname: van't Hof
authors.forename: Hidde L
authors.initials: HL
authors.lastname: Ploegh
authors.forename: Enno C I
authors.initials: EC
authors.lastname: Veerman
mesh_terms: D000595:Amino Acid Sequence; D019881:Aminoacyltransferases / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002942:Circular Dichroism; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D054808:Histatins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D006801:Humans; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010456:Peptides, Cyclic / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D011487:Protein Conformation; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D013211:Staphylococcus aureus / Q000201:enzymology / Q000235:genetics; D014945:Wound Healing / Q000187:drug effects
chemical_list: D001426:Bacterial Proteins; D054808:Histatins; D010456:Peptides, Cyclic; D011994:Recombinant Proteins; C068074:HTN1 protein, human; D019881:Aminoacyltransferases; C120542:sortase A; D003546:Cysteine Endopeptidases