Hr. Badr et al., PURIFICATION AND CHARACTERIZATION OF SUCROSE ALPHA-GLUCOHYDROLASE (INVERTASE) FROM THE HYPERTHERMOPHILIC ARCHAEON PYROCOCCUS-FURIOSUS, Systematic and applied microbiology, 17(1), 1994, pp. 1-6
Sucrose alpha-glucohydrolase (or invertase) catalyzes the hydrolysis o
f sucrose to fructose and glucose. The activity was detected in the cy
toplasmic fraction of cell-free extracts of the hyperthermophilic arch
aeon (archaebacterium), Pyrococcus furiosus, an organism that grows op
timally at 100-degrees-C. The enzyme was purified 570-fold to electrop
horetic homogeneity. It is a monomeric protein of M(r) 114,000. It exh
ibited optimum activity, as measured by the production of glucose from
sucrose, at pH 5.5 and temperatures between 100 and 117-degrees-C. Wh
ile maltose was also hydrolyzed by this enzyme, no activity could be d
etected towards starch. Kinetic analyses at 95-degrees-C established K
(m) and V(max) values for sucrose hydrolysis of 0.12 M, and 130 mumole
min-1 mg-1, respectively. Classical competitive inhibition was observ
ed in the presence of glucose or fructose with inhibition constants (K
(i)) of 0.55 mM and 50 mM, respectively. Nevertheless, the enzyme was
capable of partially hydrolyzing sucrose at concentrations up to 2.0 M
at 90-degrees-C. However, the P. furiosus enzyme is by far the most t
hermostable invertase yet reported, exhibiting a half life of almost 2
days at 95-degrees-C.