Cloning and molecular modelling of pectin degrading glycosyl hydrolase of family 28 from soil metagenomic library.

Western Ghats of India is recognized as one of
the 12 mega diversity regions of the world and is the hot spot
for unrevealed microbial diversity. To explore the diversity
of polysaccharide degrading enzymes in that region, metagenomic
library was constructed from forest soil of
Southern Western Ghats region. Nine pectinolytic clones
with the ability to degrade citrus pectin were isolated based
on function based screening of the library. Sequence analysis
of pg_4 clone containing revealed that it contained GH
family 28 domain (pfam00295) belonging to polygalacturonase
superfamily (PLN03003). Its amino acid sequence
analysis showed 25–55 % identity to the other well-characterized
polygalacturonases. Molecular modeling of pg_4
revealed that it comprised of three right handed-parallel b
sheets, one anti-parallel b sheet and one a helix with three
conserved catalytic residue D 2263, D 284-85 and H 312 at
the C terminal end. The enzyme characterized was able to
hydrolyze both apple and citrus pectin with Km values of
1.685 and 1.542 mg ml-1 and retained more that 80 % of
activity at pH 5–9 and temperature 20–60 �C.