(548e) Discovery of Thermostable Cellobiohydrolase from Mesophilic Marine Aspergillus Niger | AIChE

(548e) Discovery of Thermostable Cellobiohydrolase from Mesophilic Marine Aspergillus Niger

Authors 

Cai, L. N. - Presenter, Zhejiang University
Lin, D. Q., Zhejiang University
Yao, S. J., Zhejiang University
Li, L., Zhejiang University
The thermostable cellulases make the utilization of lignocellulosic biomass stable and efficient at high temperature. Quantities of thermostable cellulases have been identified from thermophiles or hyperthermophiles. However, few studies focused on discovery of thermostable cellulases from mesophiles. Here, a thermostable cellobiohydrolase was obtained from mesophilic marine Aspergillus niger.

Two cellobiohydrolase genes (cel71 and cel72) were cloned from marine A. niger genome. Then the expression cassettes PgpdA- cel71/cel72-Tcbh1 were inserted into marine A. niger genome for constitutive homologous expression. Two cellobiohydrolases (AnCel7I and AnCel7II) were purified to homogeneity through ultrafiltration and anion exchange chromatography. Subsequent enzyme assay revealed their optimal temperature was 55 °C, the optimal pH of AnCel7I and AnCel7II was 2.5-4.0 and 3.0-3.5, respectively. In comparison, AnCel7I was acidophilic and had a broader pH optimum, which was potential in industrial application. Notably, AnCel7II exhibited an excellent thermostability at the range of pH 5.0-8.0. Specially, at pH 5.0, its half-lives at 90 and 100 °C were 45 and 20 min, which could rival the thermostable cellobiohydrolases from thermophiles or hyperthermophiles. Analysis to amino acid residue composition revealed the percentage of polar amino acid residues in AnCel7II was 20.9% compared with 17.5% in AnCel7I. The result of molecular dynamics simulation revealed the number of salt bridges in AnCel7II was 43 compared with 25 in AnCel7I. In summary, the more polar amino acid residues in AnCel7II engendered more salt bridges, which probably made the structure more solid and stable.

Key words: cellobiohydrolase; thermostability; mesophilic marine Aspergillus niger; constitutive homologous expression; salt bridges