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Kinetic analysis of β‐Galactosidase activity from Thermococcus kodakarensis KOD1, an extreme thermophile
Author(s) -
Hwa KuoYuan,
Subramani Boopathi,
Shen Santai,
Lee Yu-May
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.963.5
Subject(s) - thermophile , hydrolysis , enzyme , thermococcus , lactose , biochemistry , chemistry , enzyme assay , extreme environment , substrate (aquarium) , chromatography , biology , bacteria , genetics , ecology , archaea , gene
The enzyme β‐galactosidase catalyzes the hydrolysis of lactose into glucose and galactose. An extracellular β‐galactosidase of 489‐amino acid length and 57‐kDa molecular weight monomer was isolated, expressed and purified from an extreme thermophile, Thermococcus kodakarensis KOD1. Here, the enzyme kinetic parameters of the β‐Galactosidase were analyzed through the hydrolysis of the yellow chromogenic substrate, chlorophenol red β‐D‐galactopyranoside (CPRG). Ad, the red colored chlorophenol red product was measured. The optimum pH and optimum temperature for the enzyme activity were 6.5 and 100°C. The maximum enzyme activity was retained even at 110°C, the highest temperature used in the study. The kinetic parameters K m and V max of the purified enzyme were 1.4mM and 2.5 U/mg for CPRG at the optimum pH and temperature conditions. The enzyme could not be inhibited with glucosidase inhibitors. The research was supported by the National Taipei University of Technology to KYH (NTUT 100‐140‐01, NTUT 99‐140‐04) and by Academia Sinica to YML.