Biochemical characterisation and genetic analysis of aureocin A53, a new, atypical bacteriocin from Staphylococcus aureus.
Aureocin A53 is produced by Staphylococcus aureus A53. It is encoded on a 10.4 kb plasmid, pRJ9, and is active against Listeria monocytogenes. Aureocin A53 is a highly cationic 51-residue peptide containing ten lysine and five tryptophan residues. Aureocin A53 was purified to homogeneity by hydrophobic-interaction, cation-exchange, and reverse-phase chromatography. MALDI-TOF mass spectrometry yielded a molecular mass of 6012.5 Da, which was 28 Da higher than predicted from the structural gene sequence of the bacteriocin. The mass increment resulted from an N-formylmethionine residue, indicating that the aureocin A53 is synthesised and secreted without a typical bacteriocin leader sequence or sec-dependent signal peptide. The structural identity of aureocin A53 was verified by Edman sequencing after de-blocking with cyanogen bromide and extensive mass spectrometry analysis of enzymatically and laser-generated fragments. The complete sequence of pRJ9 was determined and none of the open reading frames identified in the vicinity of the structural gene aucA showed similarity to genes that are typically found in bacteriocin gene clusters. Thus, neither a dedicated protease or transporter, nor modifying enzymes and regulatory elements seemed to be involved in the production of aureocin A53. Further unique features that distinguish aureocin A53 from other peptide bacteriocins include remarkable protease stability and a defined, rigid structure in aqueous solution.
Netz
Daili Jacqueline Aguilar
DJ
Departamento de Microbiologia Geral, Instituto de Microbiologia, Universidade Federal do Rio de Janeiro, CCS, Bloco I, Cidade Universitária, 21941-590 Rio de Janeiro, RJ, Brazil.
Pohl
Regula
R
Beck-Sickinger
Annette G
AG
Selmer
Thorsten
T
Pierik
Antonio J
AJ
Bastos
Maria do Carmo de Freire
Mdo C
Sahl
Hans-Georg
HG
D000595:Amino Acid Sequence; D001430:Bacteriocins / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004251:DNA Transposable Elements / Q000235:genetics; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009154:Mutation / Q000235:genetics; D016366:Open Reading Frames / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D011487:Protein Conformation; D017421:Sequence Analysis; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D013211:Staphylococcus aureus / Q000737:chemistry / Q000235:genetics
D001430:Bacteriocins; D004251:DNA Transposable Elements