A newly-isolated strain of Pseudomonas putida (HVA-1) utilized homovan
illic acid as sole carbon and energy source. Homovanillate-grown bacte
ria oxidized homovanillate and homoprotocatechuate but monohydroxylate
d and other methoxylated phenylacetic acids were oxidized poorly; meth
oxy-substituted benzoates were not oxidized. Extracts of homovanillate
-grown cells contained homoprotocatechuate 2,3-dioxygenase but the pri
mary homovanillate-degrading enzyme could not be detected. No other me
thoxylated phenylacetic acid supported growth of the organism but vani
llate was utilized as a carbon and energy source. When homovanillate-g
rown cells were used to inoculate media containing vanillate a 26 h la
g period occurred before growth commenced. Vanillate-grown bacteria ox
idized vanillate and protocatechuate but no significant oxygen uptake
was obtained with homovanillate and other phenylacetic acid derivative
s. Analysis of pathway intermediates revealed that homovanillate-grown
bacteria produced homoprotocatechuate, formaldehyde and the ring-clea
vage product 5-carboxymethyl 2-hydroxymuconic semialdehyde (CHMS) when
incubated with homovanillate but monohydroxylated or monomethoxylated
phenylacetic acids were not detected. These results suggest that homo
vanillate is degraded directly to the ring-cleavage substrate homoprot
ocatechuate by an unstable but highly specific demethylase and then un
dergoes extradiol cleavage to CHMS. It would also appear that the upta
ke/degradatory pathways for homovanillate and vanillate in this organi
sm are entirely separate and independently controlled. If stabilizatio
n of the homovanillate demethylase can be achieved, there is potential
for exploiting the substrate specificity of this enzyme in both medic
al diagnosis and in the paper industry.