The structure of a novel psychrophilic β-mannanase enzyme from Glaciozyma antarctica PI12 yeast has been modelled and analysed in detail. To our knowledge, this is the first attempt to model a psychrophilic β-mannanase from yeast. To this end, a 3D structure of the enzyme was first predicted using a threading method because of the low sequence identity (<30%) using MODELLER9v12 and simulated using GROMACS at varying low temperatures for structure refinement. Comparisons with mesophilic and thermophilic mannanases revealed that the psychrophilic mannanase contains longer loops and shorter helices, increases in the number of aromatic and hydrophobic residues, reductions in the number of hydrogen bonds and salt bridges and numerous amino acid substitutions on the surface that increased the flexibility and its efficiency for catalytic reactions at low temperatures.
Palm kernel cake (PKC) has been largely produced in Malaysia as one of the cheap and abundant agro-waste by-products from the palm oil industry and it contains high fiber (mannan) content. The present study aimed to produce β-mannanase by Bacillus subtilis ATCC11774 via optimization of the medium composition using palm kernel cake as substrate in semi-solid fermentation. The fermentation nutrients such as PKC, peptone, yeast extract, sodium chloride, magnesium sulphate (MgSO2), initial culture pH and temperature were screened using a Plackett-Burman design. The three most significant factors identified, PKC, peptone and NaCl, were further optimized using central composite design (CCD), a response surface methodology (RSM) approach, where yeast extract and MgSO2 were fixed as a constant factor. The maximum β-mannanase activity predicted by CCD under the optimum medium composition of 16.50 g/L PKC, 19.59 g/L peptone, 3.00 g/L yeast extract, 2.72 g/L NaCl and 0.2 g/L MgSO2 was 799 U/mL. The validated β-mannanase activity was 805.12 U/mL, which was close to the predicted β-mannanas activity. As a comparison, commercial media such as nutrient broth, M9 and Luria bertani were used for the production of β-mannanase with activities achieved at 204.16 ± 9.21 U/mL, 50.32 U/mL and 88.90 U/mL, respectively. The optimized PKC fermentation medium was four times higher than nutrient broth. Hence, it could be a potential fermentation substrate for the production of β-mannanase activity by Bacillus subtilis ATCC11774.