Purification, characterization, cDNA cloning and nucleotide sequencing of a cellulase from the yellow‐spotted longicorn beetle, Psacothea hilaris

• DOI: 10.1046/j.1432-1033.2003.03735.x
• Publication Date: 2003-08-04T14:43:31Z
• Authors: Masahiro Sugimura; Hirofumi Watanabe; Nathan Lo; Hitoshi Saito
• Keywords: Coleoptera; 0303 health sciences; 03 medical and health sciences; Cellulase; Larva; Animals; Cloning, Molecular; Hydrogen-Ion Concentration; Cellulose; Gastrointestinal Contents

A cellulase (endo‐β‐1,4‐glucanase, EC 3.2.1.4) was purified from the gut of larvae yellow‐spotted longicorn beetle Psacothea hilaris by acetone precipitation and elution gels after native PAGE SDS/PAGE with activity staining. The protein formed a single band, molecular mass estimated to be 47 kDa. degraded carboxymethylcellulose (CMC), insoluble cello‐oligosaccharide (average degree polymerization 34) soluble cello‐oligosaccharides longer than cellotriose, but not crystalline cellulose or cellobiose. specific against CMC 150 µmol·min −1 ·(mg protein) . TLC analysis showed that produces cellotriose cellobiose cello‐oligosaccharides. However, glucose assay linked oxidase detected small amount glucose, productivity 0.072 optimal pH P. 5.5, close in midgut larvae. N‐terminal amino‐acid sequence determined degenerate primer designed, which enabled 975‐bp cDNA clone containing typical polyadenylation signal obtained PCR sequencing. deduced high homology members glycosyl hydrolase family 5 subfamily 2, and, addition, signature for found. Thus, this is first report arthropods.