Breaking down lignin to high-value chemicals: the conversion of lignocellulose to vanillin in a gene deletion mutant of Rhodococcus jostii RHA1.

The aromatic polymer lignin represents a possible renewable source of aromatic chemicals, if biocatalytic routes for lignin breakdown can be developed. The availability of a genome sequence for Rhodococcus jostii RHA1, a bacterium that breaks down lignin, has allowed the application of a targeted pathway engineering strategy to lignin breakdown to produce vanillin, a valuable food/flavor chemical. A gene deletion strain of R.

Pathways for degradation of lignin in bacteria and fungi.

Lignin is a heterogeneous aromatic polymer found as 10-35% of lignocellulose, found in plant cell walls. The bio-conversion of plant lignocellulose to glucose is an important part of second generation biofuel production, but the resistance of lignin to breakdown is a major obstacle in this process, hence there is considerable interest in the microbial breakdown of lignin. White-rot fungi are known to break down lignin with the aid of extracellular peroxidase and laccase enzymes.

Identification of DypB from Rhodococcus jostii RHA1 as a lignin peroxidase.

Rhodococcus jostii RHA1, a polychlorinated biphenyl-degrading soil bacterium whose genome has been sequenced, shows lignin degrading activity in two recently developed spectrophotometric assays. Bioinformatic analysis reveals two unannotated peroxidase genes present in the genome of R. jostii RHA1 with sequence similarity to open reading frames in other lignin-degrading microbes.

The emerging role for bacteria in lignin degradation and bio-product formation.

The microbial degradation of lignin has been well studied in white-rot and brown-rot fungi, but is much less well studied in bacteria. Recent published work suggests that a range of soil bacteria, often aromatic-degrading bacteria, are able to break down lignin. The enzymology of bacterial lignin breakdown is currently not well understood, but extracellular peroxidase and laccase enzymes appear to be involved.

Development of novel assays for lignin degradation: comparative analysis of bacterial and fungal lignin degraders.

Two spectrophotometric assays have been developed to monitor breakdown of the lignin component of plant lignocellulose: a continuous fluorescent assay involving fluorescently modified lignin, and a UV-vis assay involving chemically nitrated lignin. These assays have been used to analyse lignin degradation activity in bacterial and fungal lignin degraders, and to identify additional soil bacteria that show activity for lignin degradation. Two soil bacteria known to act as aromatic degraders, Pseudomonas putida and Rhodococcus sp.

Investigation of acid-base catalysis in the extradiol and intradiol catechol dioxygenase reactions using a broad specificity mutant enzyme and model chemistry.

The extradiol and intradiol catechol dioxygenase reaction mechanisms proceed via a common proximal hydroperoxide intermediate, which is processed via different Criegee 1,2-rearrangements. An R215W mutant of extradiol dioxygenase MhpB, able to produce a mixture of extradiol and intradiol cleavage products, was analysed at pH 5.2-8.