|
KMID : 0624620090420010047
|
|
BMB Reports
2009 Volume.42 No. 1 p.47 ~ p.52
|
|
|
Biochemical characterization of Alanine racemase- a spore protein produced by Bacillus anthracis
|
|
Kanodia Shivani
Agarwal Shivangi
Singh Priyanka
Agarwal Shivani
Singh Preeti
Bhatnagar Rakesh
|
|
|
Abstract
|
|
|
|
Alanine racemase catalyzes the interconversion of L-alanine and D-alanine and plays a crucial role in spore germination and cell wall biosynthesis. In this study, alanine racemase produced by Bacillus anthracis was expressed and purified as a monomer in Escherichia coli and the importance of lysine 41 in the cofactor binding octapeptide and tyrosine 270 in catalysis was evaluated. The native enzyme exhibited an apparent Km of 3 mM for L-alanine, and a Vmax of 295 μmoles/min/mg, with the optimum activity occurring at 37oC and a pH of 8-9. The activity observed in the absence of exogenous pyridoxal 5 ?-phosphate suggested that the cofactor is bound to the enzyme. Additionally, the UV-visible absorption spectra indicated that the activity was pH independece, of VV-visible absorption spectra suggests that the bound PLP exists as a protonated Schiff’s base. Furthermore, the loss of activity observed in the apoenzyme suggested that bound PLP is required for catalysis. Finally, the enzyme followed non-competitive and mixed inhibition kinetics for hydroxylamine and propionate with a Ki of 160 μM and 30 mM, respectively.
|
|
|
KEYWORD
|
|
|
Alanine racemase, Bacillus anthracis, Enantiomers, Exosporium, Spore germination
|
|
|
FullTexts / Linksout information
|
|
|
|
|
|
Listed journal information
|
|
  
|