Dataset Information

Full-length nisin immunity protein NisI from Lactococcus lactis in a lipid-free form: crystallization and X-ray analysis.

ABSTRACT: NisI is a lantibiotic-binding lipoprotein that is specific for nisin. Nisin-producing microorganisms use NisI as an immunity protein for self-protection against nisin. Here, the purification, crystallization and preliminary X-ray diffraction of full-length NisI from Lactobacillus lactis in a lipid-free form (NisI22-C) are reported. Importantly, reductive methylation of the lysine residues in NisI22-C was essential for initial crystallization. Only methylated NisI22-C crystallized. The optimized crystals of methylated NisI22-C were grown in 30-40?mM ammonium sulfate, 0.1?M sodium acetate pH 4.6, 16-18% PEG 4000 at 295?K and diffracted to 1.9?Å resolution. The crystal belonged to space group P212121, with unit-cell parameters a = 45.99, b = 76.67, c = 76.39?Å, ? = ? = ? = 90.0°. Assuming the presence of one molecule in the asymmetric unit, the estimated Matthews coefficient (VM) is 2.58?Å3?Da-1 and the estimated solvent content is 52.3%.

SUBMITTER: Jeong JH

PROVIDER: S-EPMC5505245 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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Full-length nisin immunity protein NisI from Lactococcus lactis in a lipid-free form: crystallization and X-ray analysis.

Jeong Jin Hee JH Ha Sung Chul SC

Acta crystallographica. Section F, Structural biology communications 20170617 Pt 7

NisI is a lantibiotic-binding lipoprotein that is specific for nisin. Nisin-producing microorganisms use NisI as an immunity protein for self-protection against nisin. Here, the purification, crystallization and preliminary X-ray diffraction of full-length NisI from Lactobacillus lactis in a lipid-free form (NisI22-C) are reported. Importantly, reductive methylation of the lysine residues in NisI22-C was essential for initial crystallization. Only methylated NisI22-C</ ...[more]

PMID: 28695849

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Project description:Bacteria adapt to the constantly changing environment by regulating their metabolism. The global transcriptional regulator CodY is known to regulate metabolism in low-G+C Gram-positive bacteria. Systems-level identification of its direct targets by proteome and chromatin immunoprecipitation followed by sequencing (ChIP-seq) assays have rarely been reported. Here, we identified that CodY serves as an activator or a repressor of hundreds of genes involved in nitrogen metabolism, carbohydrate metabolism, and transcription through iTRAQ proteome and ChIP-seq. Combined with the electrophoretic mobility shift assay (EMSA), apart from the genes associated with amino acid biosynthesis (ilvD, leuA, optS, ybbD, dtpT, and pepN), genes involved in cell wall synthesis (murD and ftsW) and nisin immunity (nisI) were identified as being regulated by CodY. Moreover, it was demonstrated by nisin resistance assay that CodY activated the transcription of nisI and contributed to nisin immunity. Intriguingly, CodY showed a self-regulation through binding to the motif AAAGGTGTGACAACT in the coding sequence (CDS) region of codY, as verified by DNase I footprinting assay and MEME analysis. In addition, a novel conserved AT-rich motif, AATWTTCTGACAATT, was obtained in L. lactis F44. This study provides new insights into the comprehensive CodY regulation in L. lactis by controlling metabolism, nisin immunity, and self-expression. IMPORTANCE Lactococcus lactis, a species of lactic acid bacteria (LAB) widely used in food fermentation, has been the model strain in genetic engineering, and its application has extended from food to microbial cell factories. CodY is a global regulator in low-G+C Gram-positive bacteria. Its function and direct target genes at the genome-level are little known in L. lactis. In this study, we describe the comprehensive regulation mechanism of CodY. It widely modulated the metabolism of nitrogen and carbohydrate, cell wall synthesis, and nisin immunity in L. lactis F44, and its expression level was regulated by feedback control.

Dataset Information

Full-length nisin immunity protein NisI from Lactococcus lactis in a lipid-free form: crystallization and X-ray analysis.

Publications

Full-length nisin immunity protein NisI from Lactococcus lactis in a lipid-free form: crystallization and X-ray analysis.

Similar Datasets

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