Biochemical characterization, low-resolution SAXS structure and an enzymatic cleavage pattern of BlCel48 from Bacillus licheniformis.

Economic sustainability of modern biochemical technologies for plant cell wall transformations in renewable fuels, green chemicals, and sustainable materials is considerably impacted by the elevated cost of enzymes. Therefore, there is a significant drive toward discovery and characterization of novel carbohydrate-active enzymes. Here, the BlCel48 cellulase from Bacillus licheniformis, a glycoside hydrolase family 48 member (GH48), was functionally and biochemically characterized. The enzyme is catalytically stable in a broad range of temperatures and pH conditions with its enzymatic activity at pH5.0 and 60°C. BlCel48 exhibits high hydrolytic activity against phosphoric acid swollen cellulose (PASC) and bacterial cellulose (BC) and significantly lower activity against carboxymethylcellulose (CMC). BlCel48 releases predominantly cellobiose, and also small amounts of cellotriose and cellotetraose as products from PASC hydrolysis. Small-angle X-ray scattering (SAXS) data analysis revealed a globular molecular shape and monomeric state of the enzyme in solution. Its molecular mass estimated based on SAXS data is ~77.2kDa. BlCel48 has an (αα)6-helix barrel-fold, characteristic of GH48 members. Comparative analyses of homologous sequences and structures reveal the existence of two distinct loops in BlCel48 that were not present in other structurally characterized GH48 enzymes which could have importance for the enzyme activity and specificity.