Identification and characterization of novel transglycosylating a-glucosidase from Aspergillus neoniger.

Aim: Aspergillus niger is well established for secreting a-glucosidase having
transglycosylation activity, which is used as processing aid for synthesis of
isomaltooligosaccharides. The present study focuses on identification and
characterization of a non-niger Aspergillus isolate and its gene conferring
strong transglycosylation activity.
Methods and Results: The soil isolate was identified as Aspergillus neoniger
belonging to Aspergillus section Nigri using ITS (internal transcribed spacer)
and b-tubulin analysis. The sequence analysis of gene responsible for aglucosidase synthesis revealed significant nucleotide variations when compared
to other Aspergillus species. Molecular docking studies using the homology
model revealed the presence of threonine at 694 subsite position instead of
asparagine as in case of A. niger’s a-glucosidase. The enzyme was purified to
several fold using DEAE Sepharose-CL6B column and on SDS-PAGE analysis,
it was found to be 145 kDa. MS/MS analysis of the purified enzyme validated
the presence of threonine at 694 position. Commercial a-glucosidase
(Transglucosidase L ‘Amano’) derived from A. niger and the a-glucosidase
from isolate were compared for transglycosylation activity using constant test
conditions. a-glucosidase from the isolate produced 27�4% higher panose when
compared to that of commercial enzyme. Moreover, the rate of secondary
hydrolysis of panose is much lower in case of the isolate’s enzyme.
Conclusions: Fungal isolate A. neoniger was characterized, and its gene
conferring a-glucosidase activity was established for strong transglycosylation
activity having higher panose yields.
Significance and Impact of the Study: To the best of our knowledge, this is
the first report to establish a variant of a-glucosidase having strong
transglycosylation activity from A. neoniger strain. We have demonstrated that
this enzyme when used as processing aid could improve panose significantly,
which is a potential prebiotic. Also, the sequence analysis established in our
studies could provide pointers for directed evolution of this enzyme to further
improve transglycosylation activity