In Vivo Detection of Low Molecular Weight Platform Chemicals and Environmental Contaminants by Genetically Encoded Biosensors.

Genetically encoded biosensor systems operating in living cells are versatile, cheap, and transferable tools for the detection and quantification of a broad range of small molecules. This review presents state-of-the-art biosensor designs and assemblies, featuring transcription factor-, riboswitch-, and enzyme-coupled devices, highly engineered fluorescent probes, and emerging two-component systems. Importantly, (bioinformatic-assisted) strategies to resolve contextual issues, which cause biosensors to miss performance criteria in vivo, are highlighted. The optimized biosensing circuits can be used to monitor chemicals of low molecular mass (<200 g mol-1) and physicochemical properties that challenge conventional chromatographical methods with high sensitivity. Examples herein include but are not limited to formaldehyde, formate, and pyruvate as immediate products from (synthetic) pathways for the fixation of carbon dioxide (CO2), industrially important derivatives like small- and medium-chain fatty acids and biofuels, as well as environmental toxins such as heavy metals or reactive oxygen and nitrogen species. Lastly, this review showcases biosensors capable of assessing the biosynthesis of platform chemicals from renewable resources, the enzymatic degradation of plastic waste, or the bioadsorption of highly toxic chemicals from the environment. These applications offer new biosensor-based manufacturing, recycling, and remediation strategies to tackle current and future environmental and socioeconomic challenges including the wastage of fossil fuels, the emission of greenhouse gases like CO2, and the pollution imposed on ecosystems and human health.

Whole-cell cascade for the preparation of enantiopure ß-O-4 aryl ether compounds with glutathione recycling.

Bacterial ß-etherases and glutathione lyases are glutathione-dependent enzymes that catalyze the selective cleavage of ß-O-4 aryl ether bonds found in lignin. Their glutathione (GSH) dependence is regarded as major limitation for their application in the production of aromatics from lignin polymer and oligomers, as stoichiometric glutathione amounts are required. Thus, recycling of the GSH cofactor by a NAD(P)H-dependent glutathione reductase was proposed previously. Herein, the use of a whole-cell catalyst was studied for efficient ß-O-4 aryl ether bond cleavage with intracellular GSH supply and recycling. After optimization of the whole-cell catalyst as well as reaction conditions, up to 5 mM lignin model substrate 2,6-methoxyphenoxy-α-veratrylglycerone (2,6-MP-VG) were efficiently converted into 2,6-methoxyphenol (2,6-MP) and veratryl glycerone (VG) without addition of external GSH. Unexpectedly, no glucose supply was required for glutathione recycling within the cells up to this substrate concentration. To demonstrate the applicability of this whole-cell approach, a whole-cell cascade combining a stereoselective ß-etherase (either LigE from Sphingobium sp. SYK-6 or LigF-NA from Novosphingobium aromaticivorans) and a glutathione lyase (LigG-TD from Thiobacillus denitrificans) was employed in the kinetic resolution of racemic 2,6-MP-VG. This way, enantiopure (S)- and (R)-2,6-MP-VG were obtained on semi-preparative scale without the need for external GSH supply.

Microbial ß-etherases and glutathione lyases for lignin valorisation in biorefineries: current state and future perspectives.

Lignin is the major aromatic biopolymer in nature, and it is considered a valuable feedstock for the future supply of aromatics. Hence, its valorisation in biorefineries is of high importance, and various chemical and enzymatic approaches for lignin depolymerisation have been reported. Among the enzymes known to act on lignin, ß-etherases offer the possibility for a selective cleavage of the ß-O-4 aryl ether bonds present in lignin. These enzymes, together with glutathione lyases, catalyse a reductive, glutathione-dependent ether bond cleavage displaying high stereospecificity. ß-Etherases and glutathione lyases both belong to the superfamily of glutathione transferases, and several structures have been solved recently. Additionally, different approaches for their application in lignin valorisation have been reported in the last years. This review gives an overview on the current knowledge on ß-etherases and glutathione lyases, their biochemical and structural features, and critically discusses their potential for application in biorefineries.