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Antimicrobial Agents and Chemotherapy, June 2005, p. 2399-2406, Vol. 49, No. 6
0066-4804/05/$08.00+0     doi:10.1128/AAC.49.6.2399-2406.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Proline Conformation-Dependent Antimicrobial Activity of a Proline-Rich Histone H1 N-Terminal Peptide Fragment Isolated from the Skin Mucus of Atlantic Salmon

Torben Lüders,¹ Gunn Alice Birkemo,^2, Jon Nissen-Meyer,² Øivind Andersen,³ and Ingolf F. Nes^1*

Laboratory of Microbial Gene Technology, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, POB 5003, N-1432 Ås, Norway,¹ Program of Biochemistry and Molecular Biology, Department of Molecular Biosciences, University of Oslo, POB 1041 Blindern, N-0316 Oslo, Norway,² Institute of Aquaculture Research, POB 5010, N-1432 Ås, Norway³

Received 23 November 2004/ Returned for modification 10 January 2005/ Accepted 24 February 2005

A 30-residue N-terminally acetylated peptide derived from the N-terminal part of histone H1 was identified as the dominant antimicrobial peptide in skin mucus from Atlantic salmon (Salmo salar). The peptide (termed salmon antimicrobial peptide [SAMP H1]) was purified to homogeneity by a combination of reversed-phase and cation-exchange chromatographies. By Edman degradation of the deacetylated peptide and by sequencing of the PCR-amplified DNA that encodes the peptide, the complete amino acid sequence was determined to be AEVAPAPAAAAPAKAPKKKAAAKPKKAGPS. The theoretical molecular weight of N-terminally acetylated SAMP H1 was calculated to be 2,836, which is the same as that determined by matrix-assisted laser desorption ionization mass spectrometry. The peptide was active against both gram-negative and -positive bacteria. The N-terminal acetyl group was not necessary for activity since deacetylation did not reduce the activity. A synthetic peptide whose sequence was identical to that of the isolated fragment was initially inactive but could be activated by binding it to a cation-exchange column. Treatment of the synthetic peptide when it was bound to the exchange column with peptidylproline cis-trans-isomerase increased the amount of active peptide, indicating that isomerization of the proline peptide bond(s) was necessary for activation of the synthetic peptide. Comparison of the active and inactive forms by circular dichroism and chromatographic analyses suggests that the active form, both the natural and the synthetic forms, is more structured, condensed, and rigid than the inactive form, which has a more nonstructured conformation. This work shows for the first time the importance of proline isomers in the activity of an antimicrobial peptide.

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* Corresponding author. Mailing address: Laboratory of Microbial Gene Technology, Department of Chemistry, Biotechnology, and Food Science, Norwegian University of Life Sciences, POB 5003, N-1432 Ås, Norway. Phone: (47) 64 94 94 71. Fax: (47) 64 94 14 65. E-mail: ingolf.nes{at}umb.no.

Present address: Teagasc, Biotechnology Centre, Moorepark, Fermoy, County Cork, Ireland.

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Antimicrobial Agents and Chemotherapy, June 2005, p. 2399-2406, Vol. 49, No. 6
0066-4804/05/$08.00+0     doi:10.1128/AAC.49.6.2399-2406.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

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