The Metabolism of Benzoate and Methylbenzoates via the meta‐Cleavage Pathway by Pseudomonas arvilla mt‐2

Abstract

1. Pseudomonas arvilla mt-2 grows a t the expense of benzoate, m-toluate (3-methylbenzoate) and p-toluate (4-methylbenzoate), but not o-toluate (2-methylbenzoate). Under various conditions compounds with spectra characteristic of muconic semialdehydes can be made to accumulate in the media. These spectra indicate that benzoate is metabolised through catechol and 2-hydroxymuconic semialdehyde (2-hydroxy-6-oxohexa-2,4-dienoate), m-toluate through 3-methylcatechol and 2-hydroxy-6-oxohepta-2,4-dienoate and p-toluate through 4-methylcatechol and 2-hydroxy-5-methylmuconic semialdehyde (2-hydroxy-5-methyl-6-oxohexa-2,4-dienoate).2. Freshly harvested cells grown on all three substrates only consume oxygen a t a significant rate when presented with the growth substrates themselves and catechol and the methylcatechols, but not with salicylate, protocatechuate or phenol or the cresols.3. Extracts of cells grown on these substrates contain high induced levels of the suite of meta cleavage enzymes, including both the hydrolytic branch and the 4-oxalocrotonate branch.4. The substrate specificity of the early enzymes of the pathway suggests that only one nonspecific enzyme expresses each activity in cell-free extracts and that it is nonspecifically induced during growth on all three carbon sources. 5.It is suggested that the metabolic role of the hydrolytic branch of the pathway is the assimilation of 3-methylcatechol and its precursor, and that of the 4-oxalocrotonate branch is to assimilate catechol and 4-methylcatechol and their precursors.6. o-Toluate is not metabolised because it is neither a substrate for the benzoate oxidase system nor an inducer of any of the meta pathway enzymes.7 . It appears that benzoate and m-and p-toluates are the substrate inducers of the meta pathway enzymes in this organism. The ortho pathway is induced on incubating cells with catechol and the first inducer appears to be &,cis-muconate or possibly catechol itself.The coexistence of two enzymes able to degrade the meta cleavage product of catechol, 2-hydroxymuconic semialdehyde, was first demonstrated in cell-free extracts of both benzoate-grown Azotobacter species [l] and a naphthalene-grown Pseudomonas strain NCIB 9816 [2]. Sala-Trepat and Evans [3] elucidated the two resulting pathways, which after diverging a t 2-hydroxymuconic semialdehyde converge again a t 2-oxopent-4-enoate ; they concluded that in Azotobacter only the 4-oxalocrotonate branch, involving as the first step the NADf-dependent dehydrogenation of 2-hydroxymuconic semialdehyde, was of any metabolic significance since the 2-hydroxymuconic semialdehyde hydrolase was present Enzymes. Catechol 1,2-oxygenase or catechol: oxygen 1,2-oxidoreductase (EC 1.13.1.1); catechol 2,3-oxygenase or catecho1:oxygen 2,3-oxidoredoctase (EC 1.13.1.2); NAD nucleosidase or NAD glycohydrolase (EC 3.2.2.6). 21 Eur. J. Blochem., Vol.28 in only very low uninducible levels. I n NCIB 9816 all the enzymes of the 4-oxalocrotonate branch are present [4] and again it has been proposed that catechol is metab...

The Metabolism of Aromatic Ring Fission Products by Bacillus stearothermophilus Strain IC3

“…The ketone product of 3-methylcatechol M g cleavage, HOD, was not a substrate for the dehydrogenase. These results are similar to those observed with the dehydrogenase from several Azotobacter species (Sala-Trepat & Evans, 1971 a, b) and P. putida mt-2 (Murray et al, 1972).…”

“…1. Similar pathways have been delineated in pseudornonads (Murray et al, 1972;Hopper & Taylor, 1975) In strain IC3, two different enzymes appear to be responsible for the degradation of 2-hydroxymuconic semialdehyde and its S-methylated derivative. One of these enzymes, which is not NAD dependent, gives formate as a reaction product.…”

The Plasmid-coded Metabolism of Naphthalene and 2-Methylnaphthalene in Pseudomonas Strains: Phenotypic Changes Correlated with Structural Modification of the Plasmid pWW60-1

“…(a) C120 and MLE, the first two enzymes of the ortho sequence, are induced by growth on naphthalene whereas the corresponding meta pathway enzymes, C230, HMSH and HMSD, although detectable, are at low levels and are not significantly induced. (b) Paw701 does not grow on 2-methylnaphthalene, contrary to what might be expected were it a strain with a functional meta pathway, given the well documented nonspecificity of that pathway (Bayly & Dagley, 1969;Murray et al, 1972): 2-methylnaphthalene and 4-methylsalicylate are only partially metabolized by PaW701, through 4-methylcatechol and its meta ring fission product to an unidentified brown product which is not metabolized further, indicating that the meta pathway is not completely functional. Strain Paw701 can however acquire the ability to grow on 2-methylnaphthalene by mutation in one of two ways.…”

Modified ortho-cleavage pathway in Alcaligenes eutrophus JMP134 for the degradation of 4-methylcatechol

“…Microbial degrad ation of methyl-substituted aro matics normally proceeds via meta-cleavage pathways with methylcatechols as key intermediates [2][3][4]. A problem of practical concern during waste water treatment is the degradation of methylaromatics in the presence of chloroaromatics.…”