The importance of the leader sequence for directing lanthionine formation in lacticin 481.
Lantibiotics are post-translationally modified peptide antimicrobial agents that are synthesized with an N-terminal leader sequence and a C-terminal propeptide. Their maturation involves enzymatic dehydration of Ser and Thr residues in the precursor peptide to generate unsaturated amino acids, which react intramolecularly with nearby cysteines to form cyclic thioethers termed lanthionines and methyllanthionines. The role of the leader peptide in lantibiotic biosynthesis has been subject to much speculation. In this study, mutations of conserved residues in the leader sequence of the precursor peptide for lacticin 481 (LctA) did not inhibit dehydration and cyclization by lacticin 481 synthetase (LctM) showing that not one specific residue is essential for these transformations. These amino acids may therefore be conserved in the leader sequence of class II lantibiotics to direct other biosynthetic events, such as proteolysis of the leader peptide or transport of the active compound outside the cell. However, introduction of Pro residues into the leader peptide strongly affected the efficiency of dehydration, consistent with recognition of the secondary structure of the leader peptide by the synthetase. Furthermore, the presence of a hydrophobic residue at the position of Leu-7 appears important for enzymatic processing. Based on the data in this work and previous studies, a model for the interaction of LctM with LctA is proposed. The current study also showcases the ability to prepare other lantibiotics in the class II lacticin 481 family, including nukacin ISK-1, mutacin II, and ruminococcin A using the lacticin 481 synthetase. Surprisingly, a conserved Glu located in a ring that appears conserved in many class II lantibiotics, including those not belonging to the lacticin 481 subgroup, is not essential for antimicrobial activity of lacticin 481.
Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, Illinois 61801, USA.
Gregory C
GC
Patton
Moushumi
M
Paul
Lisa E
LE
Cooper
Champak
C
Chatterjee
Wilfred A
WA
van der Donk
D020121:5' Untranslated Regions / Q000235:genetics*; D000409:Alanine / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism; D000595:Amino Acid Sequence; D000613:Aminobutyrates / Q000378:metabolism; D001430:Bacteriocins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017124:Conserved Sequence; D017931:DNA Primers; D004798:Enzymes / Q000378:metabolism; D015870:Gene Expression; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D016415:Sequence Alignment; D013440:Sulfides
D020121:5' Untranslated Regions; D000613:Aminobutyrates; D001430:Bacteriocins; D017931:DNA Primers; D004798:Enzymes; C481644:LctM protein, Lactococcus lactis; D013440:Sulfides; C082508:lacticin 481; C088694:dehydrobutyrine; C015102:dehydroalanine; C001520:lanthionine; D000409:Alanine