| dc.description.abstract | The specific activities of C23Os induced by 3 isomers of cresol in crude cell extracts from P. putida SH1 was determined towards to catechol and and catabolic derivatives, 3-methylcatechol, 4- methylcatechol and 4-chrolocatechol. It showed the same substrate specificity, suggesting this strain converts o-, m-, and p-cresol to catechol probably by the same C23O.
The thermal effect on C23O was then observed by the stability of activity as well as conformation change at various temperatures. The optimally catalytic temperature of C23Onap(SH1) was at 50℃ in 20 mM phosphate buffer, pH 7.5 but the half-life of C23Onap(SH1) at this temperature was 30 min under aerobic condition. The half-life of another C23O, C23Onap(NCIB9816-4) isolated from a known naphthalene-degrading bacterium, P. putida NCIB9816-4, was 4 hour under the same condition. It indicated that C23Onap(NCIB9816-4) was more stable than C23Onap(SH1). The structural conformation of the enzymes in solution was analyzed by using small-angle neutron scattering (SANS) technique. SANS measurements revealed distinct changes on the size of the C23Onap(SH1) between 57 and 80°C, where the size can not be restored even the temperature was then reduced. At 80°C, the size of the enzyme becomes more than twice of its native one. Accordingly, the enzyme starts to denature at 57°C and the structure was destroyed as temperature reaches 80°C. Another enzyme, C23Onap(NCIB9816-4), was also studied under the same experimental conditions. This enzyme shows slightly higher heat stability, about 5°C, in both catalytic activity and conformation.
In the meantime, the factors influencing thermal stability, including Fe2+, acetone, ethanol, and starch, on C23O were further investigated by using C23O purified from p. putida mt-2. | en_US |