Purification, biochemical and secondary structural characterisation of β-mannanase from Lactobacillus casei HDS-01...

A 36.7 kDa extracellular mannanase was originally purified from a generally regarded as safe (GRAS) lactic acid bacteria (LAB) strain Lactobacillus casei HDS-01. The LC-GC/GC determined that HDS-01 mannanase contained 338 amino acid residues. This protein belonged to glycoside hydrolase (GH) 26 Family and shared high similarity (94%) but low coverage (54%) with mannanase from Bacillus licheniformis (GQ859489.1). Circular dichroism (CD) illustrated that the proportion of α + β structural elements closely related to enzyme activity and were both negatively influenced by temperature. Fourier-transform infrared (FT-IR) showed the existence of typical protein bonds such as CO, CC, NH, CCH and CC. The CD and FT-IR collectively suggested HDS-01 mannanase a β-conformation rich (60-70%) protein. The optimal reaction condition of HDS-01 mannanase was 40 °C and pH 5.0. More than 50% activity was maintained after 2 h incubation under the condition of pH value 5.0-7.0 and 30-60 °C. This enzyme was strongly activated by Mn²⁺ and exhibited a high affinity for locust bean gum (K_m = 2.68 ± 0.02 mg mL^-1, V_max = 400.03 ± 1.22 μmol mL^-1·min^-1). The HDS-01 mannanase clarified fruit juices more efficiently than commercial mannanase. These results promised HDS-01 mannanase a bio-safe addictive used in various especially food grade industrial fields.