Open AccessFunctional Characterization of an Extreme Thermophilic Class II Fructose‐1,6‐Bisphosphate Aldolase
Author(s) -
Montigny Chantal,
Sygusch Jurgen
Publication year - 1996
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1996.0243t.x
Subject(s) - aldolase a , thermus aquaticus , aldolase b , fructose bisphosphate aldolase , thermus , thermophile , fructose , chemistry , enzyme , biochemistry , chromatography , stereochemistry
Fructose‐1,6‐bisphosphate aldolase activity has been isolated and purified to homogeneity from the extreme thermophile eubacteria Thermus aquaticus. The homogeneous enzyme is a class II aldolase as fructose‐1,6‐bisphosphate cleavage activity was strongly inhibited by EDTA, and activated by Co 2+ metal ion. Taq aldolase is a stable tetramer with estimated molecular mass of 165 kDa. The enzyme is thermostable and is not inactived after heating at 90°C for 2 h but looses 80% of activity after 1 h at 97°C. The pH profile corresponding to maximal aldolase activity is displaced to more acidic values compared to other class II aldolases. Enzyme activation by both detergents and alcohols and chromatographic behaviour on hydrophobic stationary phases is consistent with presence of hydrophobic surface regions on the soluble enzyme. Kinetic behaviour of T. aquaticus aldolase at high fructose‐l, 6‐bisphosphate concentrations indicates significant negative cooperativity. The Taq aldolase NH 2 ‐terminal sequence was determined and compared with available sequences from other class II aldolases. Significant sequence similarity was found between Tag aldolase and the thermostable aldolase from Bacillus stearothermophilus.