Study On Biochemical Properties Of Glycerophosphate Oxidase From Enterococcus Faecalis

The progress of rapid identification of pathogens by means of bacterial 16S rDNA gene PCR was reviewed.Since 16S rRNA gene existed in all bacteria and showed high degree of conservation,PCR with primers based on conserved region of 16S rDNA gene maked rapid identification of pathogens possible.The bacterium which could produce L-α-Glycerophosphate oxidase in my lab. was identified by means of bacterial 16S rDNA gene PCR. The L-α-glycerophosphate oxidase from Enterococcus faecalis was purified by 40%～80% ammonium sulfate precipitation or PEG/(NH₄)₂SO₄ double aqueous phases sysytem,column chromatographies on Q-Sepharose F·F,Phenyl-Sepharose CL-4B,Chelating Sepharose F·F and Sephacryl S-200HR.The purified L-α-Glycerophosphate oxidase migrated as a single protein band on reduced/non-reduced SDS-PAGE.The results showed that molecular weight of single peptide chain was 67.6kDa.The native molecular weight was estimated to be about 121 kDa by HPLC.So the structure of L-α-Glycerophosphate oxidase was consisted of two subunits with identical molecular weight.The optimum temperature and optimum pH for enzyme action were 35℃and pH8.0 respectively.The K_m of L-α-Glycerophosphate oxidase was 3.22×10^-2mol/L to L-a-Glycerophosphate disodium salt.Otherwise, some metal ions,some organic compounds and contaminant on the enzyme activity were studied.