Characterization and functional analysis of atl, a novel gene encoding autolysin in Streptococcus suis.
Streptococcus suis serotype 2 (S. suis 2) is an important swine and human pathogen responsible for septicemia and meningitis. A novel gene, designated atl and encoding a major autolysin of S. suis 2 virulent strain HA9801, was identified and characterized in this study. The Atl protein contains 1,025 amino acids with a predicted molecular mass of 113 kDa and has a conserved N-acetylmuramoyl-l-alanine amidase domain. Recombinant Atl was expressed in Escherichia coli, and its bacteriolytic and fibronectin-binding activities were confirmed by zymography and Western affinity blotting. Two bacteriolytic bands were shown in the sodium dodecyl sulfate extracts of HA9801, while both were absent from the atl inactivated mutant. Cell chains of the mutant strain became longer than that of the parental strain. In the autolysis assay, HA9801 decreased to 20% of the initial optical density (OD) value, while the mutant strain had almost no autolytic activity. The biofilm capacity of the atl mutant was reduced ∼30% compared to the parental strain. In the zebrafish infection model, the 50% lethal dose of the mutant strain was increased up to 5-fold. Furthermore, the adherence to HEp-2 cells of the atl mutant was 50% less than that of the parental strain. Based on the functional analysis of the recombinant Atl and observed effects of atl inactivation on HA9801, we conclude that Atl is a major autolysin of HA9801. It takes part in cell autolysis, separation of daughter cells, biofilm formation, fibronectin-binding activity, cell adhesion, and pathogenesis of HA9801.
Key Lab of Animal Bacteriology, Ministry of Agriculture, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China.
Cun-Xiang
CX
Ju
Hong-Wei
HW
Gu
Cheng-Ping
CP
Lu
D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D018441:Biofilms / Q000254:growth & development; D002448:Cell Adhesion; D004195:Disease Models, Animal; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005353:Fibronectins / Q000378:metabolism; D017353:Gene Deletion; D015870:Gene Expression; D056945:Hep G2 Cells; D006801:Humans; D007928:Lethal Dose 50; D008970:Molecular Weight; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013290:Streptococcal Infections / Q000382:microbiology; D017011:Streptococcus suis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D016019:Survival Analysis; D014774:Virulence; D015027:Zebrafish
D001426:Bacterial Proteins; D005353:Fibronectins; D011994:Recombinant Proteins; D009238:N-Acetylmuramoyl-L-alanine Amidase