Sequence analysis by NMR spectroscopy of the peptide lantibiotic epilancin K7 from Staphylococcus epidermidis K7.
The amino acid sequence of the novel lantibiotic epilancin K7 from Staphylococcus epidermidis K7 was determined by NMR spectroscopy. NMR spectroscopy was used because sequencing by conventional Edman degradation techniques was prohibited by internal sequence blocks owing to the presence of modified residues. Epilancin K7 consists of 31 residues, including two alpha,beta-didehydroalanine (one-letter code U) and two alpha,beta-didehydrobutyrine (O) residues, one lanthionine (A-S-A), two beta-methyllanthionines (A*-S-A), and six lysines. Epilancin K7 has a molecular mass of 3032 +/- 1.5 Da. The amino acid sequence of epilancin K7 was derived from both through-space dipolar proton-proton interactions and through-bond scalar proton-carbon interactions as detected by two-dimensional 1H-NOESY, 1H-ROESY and three-dimensional 1H-TOCSY-NOESY, and by two-dimensional 1H,13C-heteronuclear multiple-bond correlation spectroscopy, respectively. The sequence is as follows: [sequence: see text] The N-terminal residue X partly resembles an alanine but its exact nature is unclear. The organization of the sulfide-bridge-containing (beta-methyl-)lanthionines was revealed by 1H-NMR and 1H,13C-NMR spectroscopy. Epilancin K7 has a linear structure and a high positive net charge, and therefore is classified as a type-A lantibiotic. NMR analysis of a degraded though still active form of epilancin K7 showed that two N-terminal residues of epilancin K7 were missing, owing to decomposition at the alpha,beta-didehydro alanine at position 3; it was called the epilancin K7-(3-31)-peptide (peptide fragment of epilancin K7 consisting of positions 3-31). The usefulness of three-dimensional 1H-TOCSY-NOESY, and two-dimensional 1H,13C-heteronuclear multiple-bond correlation spectroscopy at natural abundance for the study of (modified) polypeptides is demonstrated.
van de Kamp
M
M
Nijmegen SON Research Centre, Laboratory of Biophysical Chemistry, University of Nijmegen, The Netherlands.
Horstink
L M
LM
van den Hooven
H W
HW
Konings
R N
RN
Hilbers
C W
CW
Frey
A
A
Sahl
H G
HG
Metzger
J W
JW
van de Ven
F J
FJ
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000302:isolation & purification; D001430:Bacteriocins; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010455:Peptides; D013212:Staphylococcus epidermidis / Q000737:chemistry* / Q000235:genetics
D000900:Anti-Bacterial Agents; D001430:Bacteriocins; D010455:Peptides; C092251:epilancin K7