NisT, the transporter of the lantibiotic nisin, can transport fully modified, dehydrated, and unmodified prenisin and fusions of the leader peptide with non-lantibiotic peptides.
Lantibiotics are lanthionine-containing peptide antibiotics. Nisin, encoded by nisA, is a pentacyclic lantibiotic produced by some Lactococcus lactis strains. Its thioether rings are posttranslationally introduced by a membrane-bound enzyme complex. This complex is composed of three enzymes: NisB, which dehydrates serines and threonines; NisC, which couples these dehydrated residues to cysteines, thus forming thioether rings; and the transporter NisT. We followed the activity of various combinations of the nisin enzymes by measuring export of secreted peptides using antibodies against the leader peptide and mass spectroscopy for detection. L. lactis expressing the nisABTC genes efficiently produced fully posttranslationally modified prenisin. Strikingly, L. lactis expressing the nisBT genes could produce dehydrated prenisin without thioether rings and a dehydrated form of a non-lantibiotic peptide. In the absence of the biosynthetic NisBC enzymes, the NisT transporter was capable of excreting unmodified prenisin and fusions of the leader peptide with non-lantibiotic peptides. Our data show that NisT specifies a broad spectrum (poly)peptide transporter that can function either in conjunction with or independently from the biosynthetic genes. NisT secretes both unmodified and partially or fully posttranslationally modified forms of prenisin and non-lantibiotic peptides. These results open the way for efficient production of a wide range of peptides with increased stability or novel bioactivities.
Kuipers
Anneke
A
BiOMade Technology Foundation, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
de Boef
Esther
E
Rink
Rick
R
Fekken
Susan
S
Kluskens
Leon D
LD
Driessen
Arnold J M
AJ
Leenhouts
Kees
K
Kuipers
Oscar P
OP
Moll
Gert N
GN
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism / Q000502:physiology*; D001692:Biological Transport; D003001:Cloning, Molecular; D003545:Cysteine / Q000378:metabolism; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000502:physiology*; D008969:Molecular Sequence Data; D009561:Nisin / Q000737:chemistry / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011498:Protein Precursors / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C079126:NisB protein, Lactococcus lactis; C094910:NisT protein, Lactococcus lactis; D010455:Peptides; D011498:Protein Precursors; D021382:Protein Sorting Signals; D009561:Nisin; D003545:Cysteine