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Structural and functional analyses of a novel manganese-catalase from Bacillus subtilis R5

Abeera Shaeer, Mehwish Aslam, Naeem Rashid
International journal of biological macromolecules 2021 v.180 pp. 222-233
Bacillus subtilis, Escherichia coli, absorption, active sites, bacteria, calcium, catalase, genes, half life, hydrogen peroxide, molecular weight, oxygen, pH, sodium azide
Catalases catalyze the decomposition of hydrogen peroxide into water and oxygen. Limited reports are available on characterization of manganese-catalases. We describe here molecular cloning and expression in Escherichia coli of a putative manganese-catalase gene from mesophilic bacterium, Bacillus subtilis R5. The gene product, CatBₛᵤ, produced as a soluble protein, was purified to apparent homogeneity and biochemically characterized. The absorption spectra and nonsignificant inhibition by sodium azide indicated that it is a manganese-catalase. The protein was in homohexameric form in solution, with a subunit molecular weight of 30 kDa, containing ~2 Mn²⁺ and ~1 Ca²⁺ per subunit. CatBₛᵤ showed highest activity at pH 8.0 and 55 °C. It was found to be highly active with a specific activity of 25,290 μmol min⁻¹ mg⁻¹ and apparent Kₘ and kcₐₜ values of 98 mM and 1.27 × 10⁴ s⁻¹ subunit⁻¹, respectively. Although from a mesophilic source, it exhibited a half-life of 2 h at 80 °C. Furthermore, the active site and metal binding residues in CatBₛᵤ were predicted by homology modelling and molecular docking. To the best of our knowledge, this is the first characterization of a manganese-catalase from genus Bacillus.