Molecular cloning, expression and characterization of a novel feruloyl esterase enzyme from the symbionts of termite(Coptotermes formosanus) gut.
Termites play an important role in the degradation of dead plant materials and have acquired endogenous and symbiotic cellulose digestion capabilities. The feruloyl esterase enzyme (FAE) gene amplified from the metagenomic DNA of Coptotermes formosanus gut was cloned in the TA cloning vector and subcloned into a pET32a expression vector. The Ft3-7 gene has 84% sequence identity with Clostridium saccharolyticum and shows amino acid sequence identity with predicted xylanase/chitin deacetylase and endo-1,4-beta-xylanase. The sequence analysis reveals that probably Ft3-7 could be a new gene and that its molecular mass was 18.5 kDa. The activity of the recombinant enzyme (Ft3-7) produced in Escherichia coli (E.coli) was 21.4 U with substrate ethyl ferulate and its specific activity was 24.6 U/mg protein. The optimum pH and temperature for enzyme activity were 7.0 and 37oC, respectively. The substrate utilization preferences and sequence similarity of the Ft3-7 place it in the type-D sub-class of FAE.
Rumen Microbiology Laboratory, Animal Nutrition Division, National Institute of Animal Nutrition and Physiology, Adugodi, Bangalore, India. chandrasekhariah.naip@gmail.com
Matam
M
Chandrasekharaiah
Appoothy
A
Thulasi
M
M
Bagath
Duvvuri Prasanna
DP
Kumar
Sunil Singh
SS
Santosh
Chenniappan
C
Palanivel
Vazhakkala Lyju
VL
Jose
K T
KT
Sampath
D000595:Amino Acid Sequence; D000818:Animals; D002265:Carboxylic Ester Hydrolases / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D041981:Gastrointestinal Tract / Q000382:microbiology; D006863:Hydrogen-Ion Concentration; D020049:Isoptera / Q000382:microbiology*; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D013696:Temperature
D002265:Carboxylic Ester Hydrolases; C070196:feruloyl esterase