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Purification and functional characterisation of an α‐ l ‐rhamnosidase from Penicillium citrinum MTCC‐3565
Author(s) -
Yadav Sarita,
Yadav Vinita,
Yadava Sudha,
Yadav Kapil D.S.
Publication year - 2012
Publication title -
international journal of food science and technology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.831
H-Index - 96
eISSN - 1365-2621
pISSN - 0950-5423
DOI - 10.1111/j.1365-2621.2012.02987.x
Subject(s) - chemistry , chromatography , naringin , rhamnose , enzyme , penicillium citrinum , substrate (aquarium) , hydrolysis , biochemistry , polysaccharide , food science , biology , ecology
Summary An extracellular α‐ l ‐rhamnosidase from Penicillium citrinum MTCC‐3565 has purified to homogeneity from its culture filtrate using ethanol precipitation and cation‐exchange chromatography on carboxymethyl cellulose. The purified enzyme gave a single protein band corresponding to molecular mass of 45.0 kDa in SDS‐PAGE analysis showing the purity of the enzyme preparation. The native PAGE analysis showed the monomeric nature of the purified enzyme. Using p‐nitrophenyl α‐ l ‐rhamnopyranoside as substrate, K m and V max values of the enzyme were 0.30 m m and 27.0 μ m min mg −1 , respectively. The k cat value was 20.1 s giving k cat / K m value of 67.0 m m s −1 for the same substrate. The pH and temperature optima of the enzyme were 8.5 and 50 °C, respectively. The activation energy for the thermal denaturation of the enzyme was 29.9 KJ mol −1 . The α‐ l ‐rhamnosidase was able to hydrolyse naringin, rutin and hesperidin and liberated l ‐rhamnose, indicating that the purified enzyme can be used for the preparation of α‐ l ‐rhamnose and pharmaceutically important compounds by derhamnosylation of natural glycosides containing terminal α‐ l ‐rhamnose. The α‐ l ‐rhamnosidase was active at the level of ethanol concentration present in wine, indicating that it can be used for improving wine aroma.