Abstract
The cellulase cocktail of marine Aspergillus niger exhibited
halo-tolerant and thermostable properties, which is of great potential
in industrial application. In order to excavate single tolerant
cellulase components from the complex cellulase cocktail, constitutive
homologous expression was employed for direct obtainment of the
endoglucanase (AnEGL). The constitutive expression system seems
as “separation factor” to be implanted into cell for conveniently
obtaining a single target enzyme. Enzymatic property study revealed that
AnEGL exhibited a property of halo-tolerance and an outstanding
thermostability in high salt environment. Significantly, its activity
increased by 29% and the half-life at 65 °C increased by 26.7-fold with
the presence of 4.5 M NaCl. Molecular dynamics (MD) simulation revealed
that Na+ and Cl- could form salt
bridges with charged residues, and then influenced the activity of loops
and the stability of substrate binding pocket, which accounted for the
halo-tolerance and thermostability. Further, site-specific mutagenesis
study proved that the residues Asp95 and Asp99 in the pocket were of
great concern for the halo-tolerance and thermostability. The
halo-tolerant and thermostable AnEGL was of great value in
lignocellulosic utilization and the conjectural mechanisms were of
referential significance for other tolerant enzymes.