Rhodanese-Fold Containing Proteins in Humans: Not Just Key Players in Sulfur Trafficking.

The Rhodanese-fold is a ubiquitous structural domain present in various protein subfamilies associated with different physiological functions or pathophysiological conditions in humans. Proteins harboring a Rhodanese domain are diverse in terms of domain architecture, with some representatives exhibiting one or several Rhodanese domains, fused or not to other structural domains. The most famous Rhodanese domains are catalytically active, thanks to an active-site loop containing an essential cysteine residue which allows for catalyzing sulfur transfer reactions involved in sulfur trafficking, hydrogen sulfide metabolism, biosynthesis of molybdenum cofactor, thio-modification of tRNAs or protein urmylation.

Aerobic Conditions and Endogenous Reactive Oxygen Species Reduce the Production of Infectious MS2 Phage by .

Most of the defective/non-infectious enteric phages and viruses that end up in wastewater originate in human feces. Some of the causes of this high level of inactivity at the host stage are unknown. There is a significant gap between how enteric phages are environmentally transmitted and how we might design molecular tools that would only detect infectious ones.

CRD Generated by pCARGHO: A New Efficient Lectin-Based Affinity Tag Method for Safe, Simple, and Low-Cost Protein Purification.

Purification of recombinant proteins remains a bottleneck for downstream processing. The authors engineered a new galectin 3 truncated form (CRD ), functionally and structurally characterized, with preserved solubility and lectinic activity. Taking advantage of these properties, the authors designed an expression vector (pCARGHO), suitable for CRD -tagged protein expression in prokaryotes.

Catalytic properties of a bacterial acylating acetaldehyde dehydrogenase: evidence for several active oligomeric states and coenzyme A activation upon binding.

Until the last decade, two unrelated aldehyde dehydrogenase (ALDH) superfamilies, i.e. the phosphorylating and non-phosphorylating superfamilies, were known to catalyze the oxidation of aldehydes to activated or non-activated acids.

Retinoic acid biosynthesis catalyzed by retinal dehydrogenases relies on a rate-limiting conformational transition associated with substrate recognition.

Retinoic acid (RA), a metabolite of vitamin A, exerts pleiotropic effects throughout life in vertebrate organisms. Thus, RA action must be tightly regulated through the coordinated action of biosynthetic and degrading enzymes. The last step of retinoic acid biosynthesis is irreversibly catalyzed by the NAD-dependent retinal dehydrogenases (RALDH), which are members of the aldehyde dehydrogenase (ALDH) superfamily.

Adenine binding mode is a key factor in triggering the early release of NADH in coenzyme A-dependent methylmalonate semialdehyde dehydrogenase.

Structural dynamics associated with cofactor binding have been shown to play key roles in the catalytic mechanism of hydrolytic NAD(P)-dependent aldehyde dehydrogenases (ALDH). By contrast, no information is available for their CoA-dependent counterparts. We present here the first crystal structure of a CoA-dependent ALDH.

Methylmalonate-semialdehyde dehydrogenase from Bacillus subtilis: substrate specificity and coenzyme A binding.

Methylmalonate-semialdehyde dehydrogenase (MSDH) belongs to the CoA-dependent aldehyde dehydrogenase subfamily. It catalyzes the NAD-dependent oxidation of methylmalonate semialdehyde (MMSA) to propionyl-CoA via the acylation and deacylation steps. MSDH is the only member of the aldehyde dehydrogenase superfamily that catalyzes a β-decarboxylation process in the deacylation step.

Stabilization and conformational isomerization of the cofactor during the catalysis in hydrolytic ALDHs.

Over the past 15 years, mechanistic and structural aspects were studied extensively for hydrolytic ALDHs. One the most striking feature of nearly all X-ray structures of binary ALDH-NAD(P)(+) complexes is the great conformational flexibility of the NMN moiety of the NAD(P)(+), in particular of the nicotinamide ring. However, the fact that the acylation step is efficient in GAPN (non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase) from Streptococcus mutans and in other hydrolytic ALDHs implies an optimal positioning of the nicotinamide ring relative to the hemithioacetal intermediate within the ternary complex to allow an efficient and stereospecific hydride transfer.

Invariant Thr244 is essential for the efficient acylation step of the non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase from Streptococcus mutans.

One of the most striking features of several X-ray structures of CoA-independent ALDHs (aldehyde dehydrogenases) in complex with NAD(P) is the conformational flexibility of the NMN moiety. However, the fact that the rate of the acylation step is high in GAPN (non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase) from Streptococcus mutans implies an optimal positioning of the nicotinamide ring relative to the hemithioacetal intermediate within the ternary GAPN complex to allow an efficient and stereospecific hydride transfer. Substitutions of serine for invariant Thr244 and alanine for Lys178 result in a drastic decrease of the efficiency of hydride transfer which becomes rate-limiting.

The first crystal structure of a thioacylenzyme intermediate in the ALDH family: new coenzyme conformation and relevance to catalysis.

Crystal structures of several members of the nonphosphorylating CoA-independent aldehyde dehydrogenase (ALDH) family have shown that the peculiar binding mode of the cofactor to the Rossmann fold results in a conformational flexibility for the nicotinamide moiety of the cofactor. This has been hypothesized to constitute an essential feature of the catalytic mechanism because the conformation of the cofactor required for the acylation step is not appropriate for the deacylation step. In the present study, the structure of a reaction intermediate of the E268A-glyceraldehyde 3-phosphate dehydrogenase (GAPN) from Streptococcus mutans, obtained by soaking the crystals of the enzyme/NADP complex with the natural substrate, is reported.

Mechanistic characterization of the MSDH (methylmalonate semialdehyde dehydrogenase) from Bacillus subtilis.

Homotetrameric MSDH (methylmalonate semialdehyde dehydrogenase) from Bacillus subtilis catalyses the NAD-dependent oxidation of MMSA (methylmalonate semialdehyde) and MSA (malonate semialdehyde) into PPCoA (propionyl-CoA) and acetyl-CoA respectively via a two-step mechanism. In the present study, a detailed mechanistic characterization of the MSDH-catalysed reaction has been carried out. The results suggest that NAD binding elicits a structural imprinting of the apoenzyme, which explains the marked lag-phase observed in the activity assay.

Flavin radicals, conformational sampling and robust design principles in interprotein electron transfer: the trimethylamine dehydrogenase-electron-transferring flavoprotein complex.

TMADH (trimethylamine dehydrogenase) is a complex iron-sulphur flavoprotein that forms a soluble electron-transfer complex with ETF (electron-transferring flavoprotein). The mechanism of electron transfer between TMADH and ETF has been studied using stopped-flow kinetic and mutagenesis methods, and more recently by X-ray crystallography. Potentiometric methods have also been used to identify key residues involved in the stabilization of the flavin radical semiquinone species in ETF.

Expression, purification, crystallization and preliminary X-ray diffraction data of methylmalonate-semialdehyde dehydrogenase from Bacillus subtilis.

Methylmalonate-semialdehyde dehydrogenase from Bacillus subtilis was cloned and overexpressed in Escherichia coli. Suitable crystals for X-ray diffraction experiments were obtained by the hanging-drop vapour-diffusion method using ammonium sulfate as precipitant. The crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 195.

Extensive conformational sampling in a ternary electron transfer complex.

Here we report the crystal structures of a ternary electron transfer complex showing extensive motion at the protein interface. This physiological complex comprises the iron-sulfur flavoprotein trimethylamine dehydrogenase and electron transferring flavoprotein (ETF) from Methylophilus methylotrophus. In addition, we report the crystal structure of free ETF.

Electron transfer and conformational change in complexes of trimethylamine dehydrogenase and electron transferring flavoprotein.

The trimethylamine dehydrogenase-electron transferring flavoprotein (TMADH.ETF) electron transfer complex has been studied by fluorescence and absorption spectroscopies. These studies indicate that a series of conformational changes occur during the assembly of the TMADH.