Biochemical characterization of Alanine racemase- a spore protein produced by Bacillus anthracis

Kanodia, Shivani ; Agarwal, Shivangi ; Singh, Priyanka ; Agarwal, Shivani ; Singh, Preeti ; Bhatnagar, Rakesh (2009) Biochemical characterization of Alanine racemase- a spore protein produced by Bacillus anthracis Journal of Biochemistry and Molecular Biology, 42 (1). pp. 47-52. ISSN 1225-8687

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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 37°C 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.

Item Type:Article
Source:Copyright of this article belongs to Springer.
Keywords:Alanine Racemase; Bacillus Anthracis; Enantiomers; Exosporium; Spore Germination
ID Code:93510
Deposited On:21 Jun 2012 09:51
Last Modified:21 Jun 2012 09:51

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