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J Mol Microbiol Biotechnol ; 27(3): 133-146, 2017.
Article in English | MEDLINE | ID: mdl-28456803

ABSTRACT

BACKGROUND: Arginine deiminase (ArcA) has been speculated to facilitate the intracellular survival of Streptococcus suis under acidic conditions. However, the physical and biological properties and function of SS2-ArcA have not yet been elucidated. METHODS: Recombinant SS2-ArcA (rSS2-ArcA) was expressed and purified using Ni-NTA affinity chromatography. Under various pH and temperature conditions, the enzymatic properties of purified rSS2-ArcA and crude native SS2-ArcA were determined. RESULTS: The SS2-arcA-deduced amino acid sequence contained a conserved catalytic triad (Cys399-His273-Glu218). The optimum temperature and pH of 47-kDa rSS2-ArcA and crude native SS2-ArcA were 42°C and pH 7.2. The rSS2-ArcA and crude native SS2-ArcA were stable for 3 h at 4 and 25°C, respectively. The pH stability and dependency tests suggested that rSS2-ArcA and crude native SS2-ArcA were functionally active in acidic conditions. The L-arginine substrate binding affinity (Km) values of rSS2-ArcA (specific activity 16.00 U/mg) and crude native SS2-ArcA (specific activity 0.23 U/mg) were 0.058 and 0.157 mM, respectively. rSS2-ArcA exhibited a weak binding affinity with the common ArcA inhibitors L-canavanine and L-NIO. Furthermore, the partial inactivation of SS2-ArcA significantly impaired the viability and growth of SS2 at pH 4.0, 6.0, and 7.5. CONCLUSIONS: This study profoundly demonstrated the involvement of ArcA enzymatic activity in S. suis survival under acidic conditions.


Subject(s)
Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Hydrolases/chemistry , Hydrolases/genetics , Serogroup , Streptococcus suis/enzymology , Streptococcus suis/genetics , Amino Acid Sequence , Arginine/metabolism , Bacterial Proteins/drug effects , Bacterial Proteins/metabolism , Base Sequence , Canavanine/antagonists & inhibitors , Cloning, Molecular , Enzyme Assays , Escherichia coli/genetics , Gene Expression Regulation, Bacterial , Genes, Bacterial , Hydrogen-Ion Concentration , Hydrolases/drug effects , Hydrolases/metabolism , Kinetics , Ornithine/analogs & derivatives , Ornithine/antagonists & inhibitors , Protein Stability , Recombinant Proteins/biosynthesis , Recombinant Proteins/isolation & purification , Sequence Alignment , Streptococcus suis/metabolism , Temperature
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