title: Characterization of ribonuclease III from Brucella.
abstract: Bacterial ribonuclease III (RNase III) is a highly conserved endonuclease, which plays pivotal roles in RNA maturation and decay pathways by cleaving double-stranded structure of RNAs. Here we cloned rncS gene from the genomic DNA of Brucella melitensis, and analyzed the cleavage properties of RNase III from Brucella. We identified Brucella-encoding small RNA (sRNA) by high-throughput sequencing and northern blot, and found that sRNA of Brucella and Homo miRNA precursor (pre-miRNA) can be bound and cleaved by B.melitensis ribonuclease III (Bm-RNase III). Cleavage activity of Bm-RNase III is bivalent metal cations- and alkaline buffer-dependent. We constructed several point mutations in Bm-RNase III, whose cleavage activity indicated that the 133th Glutamic acid residue was required for catalytic activity. Western blot revealed that Bm-RNase III was differently expressed in Brucella virulence strain 027 and vaccine strain M5-90. Collectively, our data suggest that Brucella RNase III can efficiently bind and cleave stem-loop structure of small RNA, and might participate in regulation of virulence in Brucella.
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China.
authors.forename: Chang-Xian
authors.initials: CX
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China.
authors.forename: Xian-Jin
authors.initials: XJ
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China.
authors.forename: Ke
authors.initials: K
authors.lastname: Zheng
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China.
authors.forename: Fang
authors.initials: F
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China.
authors.forename: Xu-Dong
authors.initials: XD
authors.lastname: Yang
authors.affiliation: Department of Preventive Veterinary Medicine, College of Animal Science & Technology, Shihezi University, Shihezi City 832003, China.
authors.forename: Chuang-Fu
authors.initials: CF
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China.
authors.forename: Huan-Chun
authors.initials: HC
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China. Electronic address: lzf6789@mail.hzau.edu.cn.
authors.forename: Zheng-Fei
authors.initials: ZF
authors.lastname: Liu
mesh_terms: D000595:Amino Acid Sequence / Q000235:genetics; D002002:Brucella / Q000201:enzymology* / Q000472:pathogenicity; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology; D009690:Nucleic Acid Conformation; D020871:RNA Stability / Q000235:genetics*; D043244:Ribonuclease III / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D043244:Ribonuclease III
keywords: Brucella; Cleavage; RNase III; sRNA