4-Hydroxybenzoate-coenzyme A ligase from Rhodopseudomonas palustris: purification, gene sequence, and role in anaerobic degradation.
Anaerobic metabolism of most aromatic acids is initiated by coenzyme A thioester formation. Rhodopseudomonas palustris grows well under anaerobic, phototrophic conditions with many aromatic acids, including benzoate and 4-hydroxybenzoate, as a carbon source. A coenzyme A ligase that reacts with 4-hydroxybenzoate was purified from 4-hydroxybenzoate-grown cells of R. palustris. This enzyme required MgATP, reduced coenzyme A, and 4-hydroxybenzoate, benzoate, or cyclohex-1,4-dienecarboxylate for optimal activity but also used phosphopantetheine, cyclohex-2,5-dienecarboxylate, and 4-fluorobenzoate at lower rates. The 4-hydroxybenzoate-coenzyme A ligase differed in molecular characteristics from a previously described benzoate-coenzyme A ligase from R. palustris, and the two ligases did not cross-react immunologically. The gene encoding the 4-hydroxybenzoate enzyme was cloned and sequenced. The deduced gene product showed about 20% amino acid identity with bacterial coenzyme A ligases involved in aerobic degradation of aromatic acids. An R. palustris mutant carrying a disrupted 4-hydroxybenzoate-coenzyme A ligase gene was unable to grow with 4-hydroxybenzoate under anaerobic conditions, indicating that the enzyme is essential for anaerobic degradation of this compound.
Gibson
J
J
Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853.
Dispensa
M
M
Fogg
G C
GC
Evans
D T
DT
Harwood
C S
CS
D000693:Anaerobiosis; D001483:Base Sequence; D003001:Cloning, Molecular; D003066:Coenzyme A Ligases / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D008970:Molecular Weight; D015183:Restriction Mapping; D012241:Rhodopseudomonas / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D017931:DNA Primers; C058481:4-hydroxybenzoate coenzyme A ligase; D003066:Coenzyme A Ligases