Characterization of an acid-stable catalase KatB isolated from Bacillus altitudinis SYBC hb4

Abstract

In recent decades, many microorganisms have been selected and bred for industrial production of alkali-stable catalases. Given the diversity and excellent properties of Bacillus catalases, the aim of this work was to screen potent acid-stable catalases from Bacillus strain(s). A strain with higher production of catalase activity was identified and designated as Bacillus altitudinis SYBC hb4 based on phenotypic properties, 16S rRNA and gyrB gene analyses. Its four catalase genes were cloned successfully and designated as katX, katB, katN1 and katN2, respectively. Three distinct catalases were detected by isozyme zymography; however, only one (KatB) was successfully identified by MALDI-TOF. KatB had an estimated molecular mass of 228 kDa, and consisted of four identical subunits of 57 kDa. KatB displayed optimal activity under conditions of pH 5.0, 30 °C and 25 mM H 2 O 2 . The apparent K m and V max were calculated to be 62 mM and 33.4 mol min −1 mg −1 , respectively. KatB can maintain at least 60 % relative activity after 6 h under conditions of pH 5.0 and 30 °C. Its activity and stability were higher compared to bovine liver catalase under slightly acid conditions (pH 5.0). Thus, B. altitudinis SYBC hb4 KatB might represent a potential acid-stable catalase used in acidic conditions.

Hydrogen Peroxide-Resistant CotA and YjqC of Bacillus altitudinis Spores Are a Promising Biocatalyst for Catalyzing Reduction of Sinapic Acid and Sinapine in Rapeseed Meal

“…M2015018). The strain was isolated from gall honey (the honeybee Apis cerana collected nectar from gall flowers), which was purchased from Apiary of Ba Ma Mi Yuan Co. Ltd., Bama, China [ 32 , 33 ]. According to reports, an appropriate concentration of Mn 2+ was often used for promoting Bacillus sporulation [ 34 ].…”

“…One unit of laccase activity (U) was defined as the amount of laccase required to oxidize 1 μmol ABTS per minute. Catalase activity was determined at 30°C by monitoring the decreased absorbance value of H 2 O 2 (Aladdin Chemical Reagent Co. Ltd., China) at 240 nm (ε 240 = 43.6 M -1 cm -1 ) [ 33 ]. One unit of catalase activity (U) was defined as the amount of catalase required to decompose 1 μmol H 2 O 2 per minute.…”

“…In addition, isoenzyme analyses of catalases and laccases were performed with 8% polyacrylamide gel (Sinopharm, China) electrphorsis (PAGE) under non-denaturing conditions. The activity staining for catalases was conducted using a chromogenic probe containing isoniazid and pyrocatechol (Aladdin, China) [ 33 ]. The laccases were visualized with 10 mM DMP (Sigma, China) [ 36 ].…”

“…The MALDI-TOF MS analyses of proteins were performed as described earlier [ 33 ]. In-gel digestion of proteins was conducted using the modified method described previously [ 13 ].…”

“…The resulting supernatant was the crude enzyme. The purification of CotA and YjqC was conducted using a previously described method [ 33 ]. The main steps of the purification were, in order, fractional precipitation, ion exchange and gel filtration chromatography.…”