Two Marine Desulfotomaculum spp. of Different Origin are Capable of Utilizing Acetone and Higher Ketones

Degradation of acetone and higher ketones has been described in detail for aerobic and nitrate-reducing bacteria. Among sulfate-reducing bacteria, degradation of acetone and other ketones is still an uncommon ability and has not been understood completely yet. In the present work, we show that Desulfotomaculum arcticum and Desulfotomaculum geothermicum are able to degrade acetone and butanone. Total proteomics of cell-free extracts of both organisms indicated an involvement of a thiamine diphosphate-dependent enzyme, a B12-dependent mutase, and a specific dehydrogenase during acetone degradation. Similar enzymes were recently described to be involved in acetone degradation by Desulfococcus biacutus. As there are so far only two described sulfate reducers able to degrade acetone, D. arcticum and D. geothermicum represent two further species with this capacity. All these bacteria appear to degrade acetone via the same set of enzymes and therefore via the same pathway.

Assessing the Thiamine Diphosphate Dependent Pyruvate Dehydrogenase E1 Subunit for Carboligation Reactions with Aliphatic Ketoacids

The synthetic properties of the Thiamine diphosphate (ThDP)-dependent pyruvate dehydrogenase E1 subunit from Escherichia coli (EcPDH E1) was assessed for carboligation reactions with aliphatic ketoacids. Due to its role in metabolism, EcPDH E1 was previously characterised with respect to its biochemical properties, but it was never applied for synthetic purposes. Here, we show that EcPDH E1 is a promising biocatalyst for the production of chiral α-hydroxyketones. WT EcPDH E1 shows a 180–250-fold higher catalytic efficiency towards 2-oxobutyrate or pyruvate, respectively, in comparison to engineered transketolase variants from Geobacillus stearothermophilus (TKGST). Its broad active site cleft allows for the efficient conversion of both (R)- and (S)-configured α-hydroxyaldehydes, next to linear and branched aliphatic aldehydes as acceptor substrates under kinetically controlled conditions. The alternate, thermodynamically controlled self-reaction of aliphatic aldehydes was shown to be limited to low levels of conversion, which we propose to be due to their large hydration constants. Additionally, the thermodynamically controlled approach was demonstrated to suffer from a loss of stereoselectivity, which makes it unfeasible for aliphatic substrates.

The mechanism of a one-substrate transketolase reaction

Transketolase catalyzes the transfer of a glycolaldehyde residue from ketose (the donor substrate) to aldose (the acceptor substrate). In the absence of aldose, transketolase catalyzes a one-substrate reaction that involves only ketose. The mechanism of this reaction is unknown. Here, we show that hydroxypyruvate serves as a substrate for the one-substrate reaction and, as well as with the xylulose-5-phosphate, the reaction product is erythrulose rather than glycolaldehyde. The amount of erythrulose released into the medium is equimolar to a double amount of the transformed substrate. This could only be the case if the glycol aldehyde formed by conversion of the first ketose molecule (the product of the first half reaction) remains bound to the enzyme, waiting for condensation with the second molecule of glycol aldehyde. Using mass spectrometry of catalytic intermediates and their subsequent fragmentation, we show here that interaction of the holotransketolase with hydroxypyruvate results in the equiprobable binding of the active glycolaldehyde to the thiazole ring of thiamine diphosphate and to the amino group of its aminopyrimidine ring. We also show that these two loci can accommodate simultaneously two glycolaldehyde molecules. It explains well their condensation without release into the medium, which we have shown earlier.

The problem of plants resistance to herbicides – inhibitors of acetolactate synthase

Aim. The purpose of the work is to analyze the available literature data associated with the emergence of resistance to herbicides ALS inhibitors. Results. Target resistance to herbicides ALS inhibitors is a consequence of genetic mutations due to the replacement of one amino acid with another in the enzyme polypeptide. The article deals with data on the action of herbicides ALS inhibitors on the functioning of the enzyme and its modification in the presence of mutations that predispose the resistance to herbicides ALS inhibitors. Brief description of the structural features of the binding of various classes of herbicides inhibitors ALS with the target enzyme and the modification of the cofactors (thiamine diphosphate and flavin adenine dinucleotide) is presented. Conclusions. Studies of recent decades have led to an increase in knowledge about the action characteristics of herbicides belonging to different classes of ALS inhibitors and the causes of resistance. The obtained results are the basis for better understanding of the mechanisms of resistance to herbicides and the development of ways to overcome them. Keywords: herbicides, acetolactate synthase, resistance, gene mutation.