A digestive beta-glucosidase from the salivary glands of the termite, Neotermes koshunensis (Shiraki): distribution, characterization and isolation of its precursor cDNA by 5'- and 3'-RACE amplifications with degenerate primers.
beta-Glucosidase activity [EC 3.2.1.21] was measured in the salivary glands and the gut of wood-eating termite, Neotermes koshunensis (Shiraki). 75% of the activity was detected in the salivary glands, whereas 15% of the activity was present in the hindgut, where numerous symbiotic flagellates reside. The salivary beta-glucosidase was partially purified by ion-exchange and gel filtration chromatography. The molecular weight of the salivary beta-glucosidase was 60 kDa, and the K(m) value on cellobiose was 2.5 mM. Its optimal pH was 5.6 and the activity was stable from 20 degrees C up to 45 degrees C. In addition to cellobiose, p-nitrophenyl-beta-D-fucopyranoside and laminaribiose were efficiently hydrolyzed by the salivary beta-glucosidase. Degenerate PCR using primers designed from N-terminal amino acid sequences of the salivary beta-glucosidase resulted in a cDNA fragment of 1730 bp, encoding 498 amino acids and with sequence similarity to glycosyl hydrolase family 1. Reverse-transcription (RT)-PCR showed that this beta-glucosidase is produced only in the salivary glands.
Tokuda
Gaku
G
Center of Molecular Biosciences, University of the Ryukyus, Nishihara, Okinawa 903-0213, Japan. tokuda@comb.u-ryukyu.ac.jp
Saito
Hitoshi
H
Watanabe
Hirofumi
H
D020413:3' Untranslated Regions / Q000235:genetics; D020121:5' Untranslated Regions / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D018076:DNA, Complementary; D020049:Isoptera / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012469:Salivary Glands / Q000201:enzymology*; D013379:Substrate Specificity; D001617:beta-Glucosidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D020413:3' Untranslated Regions; D020121:5' Untranslated Regions; D017931:DNA Primers; D018076:DNA, Complementary; D001617:beta-Glucosidase