Sec-mediated transport of posttranslationally dehydrated peptides in Lactococcus lactis.
Nisin is a lanthionine-containing antimicrobial peptide produced by Lactococcus lactis. Its (methyl)lanthionines are introduced by two posttranslational enzymatic steps involving the dehydratase NisB, which dehydrates serine and threonine residues, and the cyclase NisC, which couples these dehydrated residues to cysteines, yielding thioether-bridged amino acids called lanthionines. The prenisin is subsequently exported by the ABC transporter NisT and extracellularly processed by the peptidase NisP. L. lactis expressing the nisBTC genes can modify and secrete a wide range of nonlantibiotic peptides. Here we demonstrate that in the absence of NisT and NisC, the Sec pathway of L. lactis can be exploited for the secretion of dehydrated variants of therapeutic peptides. Furthermore, posttranslational modifications by NisB and NisC still occur even when the nisin leader is preceded by a Sec signal peptide or a Tat signal peptide 27 or 44 amino acids long, respectively. However, transport of fully modified prenisin via the Sec pathway is impaired. The extent of NisB-mediated dehydration could be improved by raising the intracellular concentration NisB or by modulating the export efficiency through altering the signal sequence. These data demonstrate that besides the traditional lantibiotic transporter NisT, the Sec pathway with an established broad substrate range can be utilized for the improved export of lantibiotic enzyme-modified (poly)peptides.
Kuipers
Anneke
A
BiOMaDe Technology Foundation, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
Wierenga
Jenny
J
Rink
Rick
R
Kluskens
Leon D
LD
Driessen
Arnold J M
AJ
Kuipers
Oscar P
OP
Moll
Gert N
GN
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism*; D001709:Biotechnology / Q000379:methods; D013294:Lactococcus lactis / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000378:metabolism*; D009561:Nisin / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals; D000069816:SEC Translocation Channels; D000081416:SecA Proteins
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; C079126:NisB protein, Lactococcus lactis; D010455:Peptides; D021382:Protein Sorting Signals; D000069816:SEC Translocation Channels; D009561:Nisin; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins