Biochemical Characterization of Phenylacetaldehyde Dehydrogenases from Styrene-degrading Soil Bacteria.

Four phenylacetaldehyde dehydrogenases (designated as FeaB or StyD) originating from styrene-degrading soil bacteria were biochemically investigated. In this study, we focused on the Michaelis-Menten kinetics towards the presumed native substrate phenylacetaldehyde and the obviously preferred co-substrate NAD. Furthermore, the substrate specificity on four substituted phenylacetaldehydes and the co-substrate preference were studied.

Toward Biorecycling: Isolation of a Soil Bacterium That Grows on a Polyurethane Oligomer and Monomer.

The fate of plastic waste and a sustainable use of synthetic polymers is one of the major challenges of the twenty first century. Waste valorization strategies can contribute to the solution of this problem. Besides chemical recycling, biological degradation could be a promising tool.

Catalytic Performance of a Class III Old Yellow Enzyme and Its Cysteine Variants.

Class III old yellow enzymes (OYEs) contain a conserved cysteine in their active sites. To address the role of this cysteine in OYE-mediated asymmetric synthesis, we have studied the biocatalytic properties of OYERo2a from 1CP (WT) as well as its engineered variants C25A, C25S and C25G. OYERo2a in its redox resting state (oxidized form) is irreversibly inactivated by -methylmaleimide.

Investigation of the co-metabolic transformation of 4-chlorostyrene into 4-chlorophenylacetic acid in ST.

The side-chain oxygenation of styrene is able to yield substituted phenylacetic acids from corresponding styrenes by co-metabolic transformation. This co-metabolization was investigated in ST using 4-chlorostyrene as co-substrate. It was shown that non-substituted styrene is necessary to ensure the co-metabolic process.

Detection of arsenic-binding siderophores in arsenic-tolerating Actinobacteria by a modified CAS assay.

The metalloid arsenic is highly toxic to all forms of life, and in many countries decontamination of water and soil is still required. Some bacteria have mechanisms to detoxify arsenic and can live in its presence. Actinobacteria are well known for their ability to produce a myriad of biologically-active compounds.

N-terminus determines activity and specificity of styrene monooxygenase reductases.

Styrene monooxygenases (SMOs) are two-enzyme systems that catalyze the enantioselective epoxidation of styrene to (S)-styrene oxide. The FADH co-substrate of the epoxidase component (StyA) is supplied by an NADH-dependent flavin reductase (StyB). The genome of Rhodococcus opacus 1CP encodes two SMO systems.

Heterologous production of different styrene oxide isomerases for the highly efficient synthesis of phenylacetaldehyde.

The styrene oxide isomerase (SOI, StyC) represents a key enzyme of the styrene-degrading pathway and has been discussed as promising biocatalyst during recent studies. The enzyme enables the synthesis of pure phenylacetaldehyde from styrene oxide. In this study the native as well as the corresponding codon-optimized genes of three different SOIs from Rhodococcus opacus 1CP (StyC-1CP), Sphingopyxis fribergensis Kp5.

Microbial diversity of the hypersaline and lithium-rich Salar de Uyuni, Bolivia.

Salar de Uyuni, situated in the Southwest of the Bolivian Altiplano, is the largest salt flat on Earth. Brines of this athalassohaline hypersaline environment are rich in lithium and boron. Due to the ever- increasing commodity demand, the industrial exploitation of brines for metal recovery from the world's biggest lithium reservoir is likely to increase substantially in the near future.

Characterization of Aldehyde Dehydrogenases Applying an Enzyme Assay with In Situ Formation of Phenylacetaldehydes.

Herein, different dehydrogenases (DH) were characterized by applying a novel two-step enzyme assay. We focused on the NAD(P)-dependent phenylacetaldehyde dehydrogenases because they produce industrially relevant phenylacetic acids, but they are not well studied due to limited substrate availability. The first assay step comprises a styrene oxide isomerase (440 U mg) which allows the production of pure phenylacetaldehydes (>70 mmol L) from commercially available styrene oxides.

A thermophilic-like ene-reductase originating from an acidophilic iron oxidizer.

Ene-reductases originating from extremophiles are gaining importance in the field of biocatalysis due to higher-stability properties. The genome of the acidophilic iron-oxidizing bacterium "Ferrovum" sp. JA12 was found to harbor a thermophilic-like ene-reductase (FOYE-1).

Functional characterization and stability improvement of a 'thermophilic-like' ene-reductase from Rhodococcus opacus 1CP.

Ene-reductases (ERs) are widely applied for the asymmetric synthesis of relevant industrial chemicals. A novel ER OYERo2 was found within a set of 14 putative old yellow enzymes (OYEs) obtained by genome mining of the actinobacterium Rhodococcus opacus 1CP. Multiple sequence alignment suggested that the enzyme belongs to the group of 'thermophilic-like' OYEs.

Catalytic and hydrodynamic properties of styrene monooxygenases from Rhodococcus opacus 1CP are modulated by cofactor binding.

Styrene monooxygenases (SMOs) are flavoenzymes catalyzing the epoxidation of styrene into styrene oxide. SMOs are composed of a monooxygenase (StyA) and a reductase (StyB). The latter delivers reduced FAD to StyA on the expense of NADH.

Sphingopyxis fribergensis sp. nov., a soil bacterium with the ability to degrade styrene and phenylacetic acid.

Strain Kp5.2(T) is an aerobic, Gram-negative soil bacterium that was isolated in Freiberg, Saxony, Germany. The cells were motile and rod-shaped.

Production of a recombinant membrane protein in an strain for the whole cell biosynthesis of phenylacetic acids.

The styrene oxide isomerase (SOI) represents a membrane-bound enzyme of the microbial styrene degradation pathway and has been discussed as promising biocatalyst. It catalyzes the isomerization of styrene oxide to phenylacetaldehyde. In this study a gene, which encodes the SOI of 1CP, was optimized for optimal expression in BL21(DE3) pLysS.

Co-metabolic formation of substituted phenylacetic acids by styrene-degrading bacteria.

Some soil bacteria are able to metabolize styrene via initial side-chain oxygenation. This catabolic route is of potential biotechnological relevance due to the occurrence of phenylacetic acid as a central metabolite. The styrene-degrading strains 1CP, ST, and the novel isolates sp.

Styrene oxide isomerase of Sphingopyxis sp. Kp5.2.

Styrene oxide isomerase (SOI) catalyses the isomerization of styrene oxide to phenylacetaldehyde. The enzyme is involved in the aerobic styrene catabolism via side-chain oxidation and allows the biotechnological production of flavours. Here, we reported the isolation of new styrene-degrading bacteria that allowed us to identify novel SOIs.

A mechanistic study on SMOB-ADP1: an NADH:flavin oxidoreductase of the two-component styrene monooxygenase of Acinetobacter baylyi ADP1.

Two styrene monooxygenase types, StyA/StyB and StyA1/StyA2B, have been described each consisting of an epoxidase and a reductase. A gene fusion which led to the chimeric reductase StyA2B and the occurrence in different phyla are major differences. Identification of SMOA/SMOB-ADP1 of Acinetobacter baylyi ADP1 may enlighten the gene fusion event since phylogenetic analysis indicated both proteins to be more related to StyA2B than to StyA/StyB.

Immobilization of an integral membrane protein for biotechnological phenylacetaldehyde production.

Styrene oxide isomerase (SOI) has previously been shown to be an integral membrane protein performing a highly selective, hydrolytic ring opening reaction of epoxides to yield pure aldehydes. Earlier studies had also shown a high sensitivity of SOIs toward their product phenylacetaldehyde which caused an irreversible inhibition and finally complete loss of activity at higher aldehyde concentrations. Here we report on the covalent immobilization of a styrene oxide isomerase (SOI) on SBA-15 silica carriers.

Flavin dependent monooxygenases.

Flavin-dependent monooxygenases catalyze a wide variety of chemo-, regio- and enantioselective oxygenation reactions. As such, they are involved in key biological processes ranging from catabolism, detoxification and biosynthesis, to light emission and axon guidance. Based on fold and function, flavin-dependent monooxygenases can be distributed into eight groups.

FAD C(4a)-hydroxide stabilized in a naturally fused styrene monooxygenase.

StyA2B represents a new class of styrene monooxygenases that integrates flavin-reductase and styrene-epoxidase activities into a single polypeptide. This naturally-occurring fusion protein offers new avenues for studying and engineering biotechnologically relevant enantioselective biochemical epoxidation reactions. Stopped-flow kinetic studies of StyA2B reported here identify reaction intermediates similar to those reported for the separate reductase and epoxidase components of related two-component systems.

Bench-scale study of the effect of phosphate on an aerobic iron oxidation plant for mine water treatment.

At the opencast pit Nochten acidic iron- and sulfate-rich mine waters are treated biotechnologically in a mine-water treatment plant by microbial iron oxidation. Due to the low phosphate concentration in such waters the treatment plant was simulated in bench-scale to investigate the influence of addition of potassium dihydrogen phosphate on chemical and biological parameters of the mine-water treatment. As a result of the phosphate addition the number of cells increased, which resulted in an increase of the iron oxidation rate in the reactor with phosphate addition by a factor of 1.

New cultivation medium for "Ferrovum" and Gallionella-related strains.

Since the first isolation of the well-known iron oxidizer Acidithiobacillus ferrooxidans various media and techniques have been developed to isolate new species of acidophilic iron-oxidizing bacteria. A successful strategy in many cases was the use of iFeo medium in double-layer plates with a heterotrophic strain in the underlayer. However, even with samples which had been shown by molecular techniques to be dominated by "Ferrovum myxofaciens" and Gallionella-related bacteria, these bacteria were isolated considerably less frequently than Acidithiobacillus spp.

Trehalose phosphate synthases OtsA1 and OtsA2 of Rhodococcus opacus 1CP.

Rhodococcus opacus 1CP produces trehalose dinocardiomycolates during growth on long-chained n-alkanes. Trehalose and trehalose-6-phosphate, which are synthesized via the OtsAB pathway, are probable intermediates in the biosynthesis of these biosurfactants. By molecular genetic screening for trehalose-6-phosphate synthases (TPSs and OtsAs), two chromosomal fragments of strain 1CP were obtained.

One-component styrene monooxygenases: an evolutionary view on a rare class of flavoproteins.

Styrene monooxygenases (SMOs) are catalysts for the enantioselective epoxidation of terminal alkenes. Most representatives comprise a reductase and a monooxygenase which are encoded by separate genes (styA, styB). Only six presumed self-sufficient one-component SMOs (styA2B) have previously been submitted to databases, and one has so far been characterized.

Styrene oxide isomerase of Rhodococcus opacus 1CP, a highly stable and considerably active enzyme.

Styrene oxide isomerase (SOI) is involved in peripheral styrene catabolism of bacteria and converts styrene oxide to phenylacetaldehyde. Here, we report on the identification, enrichment, and biochemical characterization of a novel representative from the actinobacterium Rhodococcus opacus 1CP. The enzyme, which is strongly induced during growth on styrene, was shown to be membrane integrated, and a convenient procedure was developed to highly enrich the protein in active form from the wild-type host.