QUANTITATIVE STRUCTURE ACTIVITY RELATIONSHIP FOR THE RATE OF CONVERSION OF C4-SUBSTITUTED CATECHOLS BY CATECHOL-1,2-DIOXYGENASE FROM PSEUDOMONAS-PUTIDA (ARVILLA) C1/
L. Ridder et al., QUANTITATIVE STRUCTURE ACTIVITY RELATIONSHIP FOR THE RATE OF CONVERSION OF C4-SUBSTITUTED CATECHOLS BY CATECHOL-1,2-DIOXYGENASE FROM PSEUDOMONAS-PUTIDA (ARVILLA) C1/, European journal of biochemistry, 257(1), 1998, pp. 92-100
The influence of various C4/C5 substituents in catechol (1,2-dihydroxy
benzene) derivatives on the overall rate of conversion by catechol-1.2
-dioxygenase from Pseudomonas putida (arvilla) C1 was investigated. Us
ing catechol. 4-methylcatechol. 4-fluorocatechol, 4-chlorocatechol. 4-
bromocatechol, 4,5-difluorocatechol and 4-chloro-5-fluorocatechol, it
could be demonstrated that substituents at the C4 and/or C5 position d
ecrease the rate of conversion, from 62% (4-methylcatechol) down to 0.
7% (4-chloro-5-fluorocatechol) of the activity with non-substituted ca
techol, The inhibition was reversible upon addition of excess catechol
for all substrates tested. This indicates that the lower activities a
re neither due to irreversible inactivation of the enzyme nor to produ
ct inhibition. Based on the reaction mechanism proposed in the literat
ure [Que, L. & Ho, R. Y. N. (1996) Chem. Rev. 96. 2606-2624], the nucl
eophilic reactivity of the catecholate was expected to be an essential
characteristic for its conversion by catechol-1,2-dioxygenase. Theref
ore, the fates of conversion were compared with calculated energies of
the highest occupied molecular orbital (E-Homo) of the substrates. A
clear quantitative relationship (R>0.97) between the In k(cai) and the
calculated electronic parameter E-HOMO was obtained. This indicates t
hat the rate-limiting step of the reaction cycle is dependent on the n
ucleophilic reactivity of the substrate and not sterically hindered by
the relatively large bromine or methyl substituents used in the prese
nt study. Possible steps in the reaction mechanism determining the ove
rall rate at 20 degrees C are discussed.