Influence of shifting positions of Ser, Thr, and Cys residues in prenisin on the efficiency of modification reactions and on the antimicrobial activities of the modified prepeptides.
Since the recent discovery that the nisin modification and transport machinery can be used to produce and modify peptides unrelated to nisin, specific questions arose concerning the specificity of the modification enzymes involved and the limits of their promiscuity with respect to the dehydration and cyclization processes. The nisin leader peptide has been postulated to fulfill a recognition and binding function required for these modifications. Here, we investigated whether the relative positions of the modifiable residues in the nisin prepeptide, with respect to the leader peptide, could influence the efficiency of their modification. We conducted a systematic study on the insertion of one to four alanines in front of either ring A or ring D to change the "reading frame" of modifiable residues, resulting in altered distance and topology of the modifiable residues relative to the leader. The insertion of N-terminal and hinge-located Ala residues had only a modest influence on the modification efficiency, demonstrating that the "phasing" of these residues relative to the leader peptide is not a critical factor in determining modification. However, in all cases, but especially with the N-terminal insertions, the antimicrobial activities of the fully modified nisin species were decreased.
Molecular Genetics Department, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.
Jacek
J
Lubelski
Wout
W
Overkamp
Leon D
LD
Kluskens
Gert N
GN
Moll
Oscar P
OP
Kuipers
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001665:Binding Sites; D003545:Cysteine / Q000378:metabolism*; D013294:Lactococcus lactis / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D008969:Molecular Sequence Data; D009561:Nisin / Q000737:chemistry* / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology*; D010957:Plasmids; D011498:Protein Precursors / Q000737:chemistry / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics / Q000502:physiology; D012694:Serine / Q000378:metabolism*; D013912:Threonine / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D010455:Peptides; D011498:Protein Precursors; D021382:Protein Sorting Signals; D009561:Nisin; D013912:Threonine; D012694:Serine; D003545:Cysteine