title: Transcriptional regulation and characteristics of a novel N-acetylmuramoyl-L-alanine amidase gene involved in Bacillus thuringiensis mother cell lysis.
abstract: In Bacillus thuringiensis, a novel N-acetylmuramoyl-L-alanine amidase gene (named cwlB) was detected, and the CwlB protein was purified and characterized. Reverse transcription-PCR (RT-PCR) results indicated that cwlB and an upstream gene (named cwlA) formed one transcriptional unit. 5' rapid amplification of cDNA ends (5'-RACE)-PCR and transcriptional fusions with the lacZ gene indicated that transcription of the operon was directed by a promoter, P(cwlA), which is located upstream from the cwlA gene and that the transcription start site is a single 5'-end nucleotide residue T located 25 nucleotides (bp) upstream from the cwlA translational start codon. Moreover, the activity of P(cwlA) was controlled by σ(K). Morphological analysis suggested that the mutation of cwlB could delay spore release compared to the timing of spore release in the wild-type strain. Western blot assay demonstrated that purified CwlB bound to the B. thuringiensis cell wall. Observations with laser confocal microscopy and a green fluorescent protein-based reporter system demonstrated that the CwlB protein localizes to the cell envelope. All results suggest that the CwlB protein is involved in mother cell lysis in B. thuringiensis.
authors.affiliation: College of Life Sciences, Northeast Agriculture University, Harbin, China.
authors.forename: Jingni
authors.initials: J
authors.lastname: Yang
authors.forename: Qi
authors.initials: Q
authors.lastname: Peng
authors.forename: Zhen
authors.initials: Z
authors.lastname: Chen
authors.forename: Chao
authors.initials: C
authors.lastname: Deng
authors.forename: Changlong
authors.initials: C
authors.lastname: Shu
authors.forename: Jie
authors.initials: J
authors.lastname: Zhang
authors.forename: Dafang
authors.initials: D
authors.lastname: Huang
authors.forename: Fuping
authors.initials: F
authors.lastname: Song
mesh_terms: D019677:Artificial Gene Fusion; D001413:Bacillus thuringiensis / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D001433:Bacteriolysis; D015153:Blotting, Western; D002473:Cell Wall / Q000378:metabolism; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014157:Transcription Factors / Q000378:metabolism; D024363:Transcription Initiation Site; D014158:Transcription, Genetic; D001616:beta-Galactosidase / Q000032:analysis / Q000235:genetics
chemical_list: D014157:Transcription Factors; C084190:sigma K; D001616:beta-Galactosidase; D009238:N-Acetylmuramoyl-L-alanine Amidase