title: Residue histidine 669 is essential for the catalytic activity of Bacillus anthracis lethal factor.
abstract: The lethal factor (LF) of Bacillus anthracis is a Zn(2+)-dependent metalloprotease which plays an important role in anthrax virulence. This study was aimed at identifying the histidine residues that are essential to the catalytic activities of LF. The site-directed mutagenesis was employed to replace the 10 histidine residues in domains II, III, and IV of LF with alanine residues, respectively. The cytotoxicity of these mutants was tested, and the results revealed that the alanine substitution for His-669 completely abolished toxicity to the lethal toxin (LT)-sensitive RAW264.7 cells. The reason for the toxicity loss was further explored. The zinc content of this LF mutant was the same as that of the wild type. Also this LF mutant retained its protective antigan (PA)-binding activity. Finally, the catalytic cleavage activity of this mutant was demonstrated to be drastically reduced. Thus, we conclude that residue His-669 is crucial to the proteolytic activity of LF.
authors.affiliation: National Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, People's Republic of China.
authors.forename: Sha
authors.initials: S
authors.lastname: Cao
authors.forename: Aizhen
authors.initials: A
authors.lastname: Guo
authors.forename: Gaobing
authors.initials: G
authors.lastname: Wu
authors.forename: Ziduo
authors.initials: Z
authors.lastname: Liu
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.forename: Chunfang
authors.initials: C
authors.lastname: Feng
authors.forename: Cheng-Cai
authors.initials: CC
authors.lastname: Zhang
authors.forename: Huanchun
authors.initials: H
authors.lastname: Chen
mesh_terms: D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics / Q000378:metabolism*; D001408:Bacillus anthracis / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D006639:Histidine / Q000737:chemistry*; D051379:Mice; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013154:Spleen / Q000473:pathology
chemical_list: D000942:Antigens, Bacterial; D001427:Bacterial Toxins; C030325:anthrax toxin; D006639:Histidine