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2,4,6-Trinitrophenol (picric acid) and 2,4-dinitrophenol were readily biodegraded by the strain Nocardioides simplex FJ2-1A. Aerobic bacterial degradation of these -electron-deficient aromatic compounds is initiated by hydrogenation at the aromatic ring. A two-component enzyme system was identified which catalyzes hydride transfer to picric acid and 2,4-dinitrophenol. Enzymatic activity was dependent on NADPH and coenzyme F 420 . The latter could be replaced by an authentic preparation of coenzyme F 420 from Methanobacterium thermoautotrophicum. One of the protein components functions as a NADPH-dependent F 420 reductase. A second component is a hydride transferase which transfers hydride from reduced coenzyme F 420 to the aromatic system of the nitrophenols. The N-terminal sequence of the F 420 reductase showed high homology with an F 420 -dependent NADP reductase found in archaea. In contrast, no N-terminal similarity to any known protein was found for the hydride-transferring enzyme.

Biodegradation tests according to Organization for Economic Cooperation and Development standard 301F(manometric respirometry test) with technical iminodisuccinate (IDS) revealed ready biodegradability for all stereoisomers of IDS. The IDS-degrading strain Agrobacterium tumefaciens BY6 was isolated from activated sludge. The strain was able to grow on each IDS isomer as well as on Fe 2؉ -, Mg 2؉ -, and Ca 2؉ -IDS complexes as the sole carbon, nitrogen, and energy source. In contrast, biodegradation of and growth on Mn 2؉ -IDS were rather scant and very slow on Cu 2؉ -IDS. Growth and turnover experiments with A. tumefaciens BY6 indicated that the isomer R,S-IDS is the preferred substrate. The IDS-degrading enzyme system isolated from this organism consists of an IDS-epimerase and a C-N lyase. The C-N lyase is stereospecific for the cleavage of R,S-IDS, generating D-aspartic acid and fumaric acid. The decisive enzyme for S,S-IDS and R,R-IDS degradation is the epimerase. It transforms S,S-IDS and R,R-IDS into R,S-IDS. Both enzymes do not require any cofactors. The two enzymes were purified and characterized, and the N-termini were sequenced. The purified lyase and also the epimerase catalyzed the transformation of alkaline earth metal-IDS complexes, while heavy metal-IDS complexes were transformed rather slowly or not at all. The observed mechanism for the complete mineralization of all IDS isomers involving an epimerase offers an interesting possibility of funneling all stereoisomers into a catabolic pathway initiated by a stereoselective lyase.

Biodegradation of 2,4,6-trinitrophenol (picric acid) byRhodococcus erythropolis HLPM-1 proceeds via initial hydrogenation of the aromatic ring system. Here we present evidence for the formation of a hydride-Meisenheimer complex (anionic ς-complex) of picric acid and its protonated form under physiological conditions. These complexes are key intermediates of denitration and productive microbial degradation of picric acid. For comparative spectroscopic identification of the hydride complex, it was necessary to synthesize this complex for the first time. Spectroscopic data revealed the initial addition of a hydride ion at position 3 of picric acid. This hydride complex readily picks up a proton at position 2, thus forming a reactive species for the elimination of nitrite. Cell extracts ofR. erythropolis HLPM-1 transform the chemically synthesized hydride complex into 2,4-dinitrophenol. Picric acid is used as the sole carbon, nitrogen, and energy source by R. erythropolisHLPM-1.