Haloalkane dehalogenases (EC 3.8.1.5) play an important role in hydrolytic degradation of halogenated compounds, resulting in a halide ion, a proton, and an alcohol. They are used in biocatalysis, bioremediation, and biosensing of environmental pollutants and also for molecular tagging in cell biology. The method of ancestral sequence reconstruction leads to prediction of sequences of ancestral enzymes allowing their experimental characterization. Based on the sequences of modern haloalkane dehalogenases from the subfamily II, the most common ancestor of thoroughly characterized enzymes LinB from Sphingobium japonicum UT26 and DmbA from Mycobacterium bovis 5033/66 was in silico predicted, recombinantly produced and structurally characterized. The ancestral enzyme AncLinB-DmbA was crystallized using the sitting-drop vapor-diffusion method, yielding rod-like crystals that diffracted X-rays to 1.5 Å resolution. Structural comparison of AncLinB-DmbA with their closely related descendants LinB and DmbA revealed some differences in overall structure and tunnel architecture. Newly prepared AncLinB-DmbA has the highest active site cavity volume and the biggest entrance radius on the main tunnel in comparison to descendant enzymes. Ancestral sequence reconstruction is a powerful technique to study molecular evolution and design robust proteins for enzyme technologies.

• Klíčová slova
• ancestral sequence reconstruction, haloalkane dehalogenase, halogenated pollutants, structural analysis,
• MeSH
• hydrolasy chemie   metabolismus   MeSH
• hydrolýza MeSH
• katalytická doména MeSH
• krystalografie rentgenová metody   MeSH
• molekulární evoluce MeSH
• molekulární modely MeSH
• Mycobacterium bovis enzymologie   MeSH
• proteinové inženýrství metody   MeSH
• sekvenční analýza proteinů metody   MeSH
• Sphingomonadaceae enzymologie   MeSH
• vazebná místa MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• haloalkane dehalogenase MeSH Prohlížeč
• hydrolasy MeSH

Fibroblast growth factor 2 (FGF2) is a signaling protein that plays a significant role in tissue development and repair. FGF2 binds to fibroblast growth factor receptors (FGFRs) alongside its co-factor heparin, which protects FGF2 from degradation. The binding between FGF2 and FGFRs induces intracellular signaling pathways such as RAS-MAPK, PI3K-AKT, and STAT. FGF2 has strong potential for application in cell culturing, wound healing, and cosmetics but the potential is severely limited by its low protein stability. The thermostable variant FGF2-STAB was constructed by computer-assisted protein engineering to overcome the natural limitation of FGF2. Previously reported characterization of FGF2-STAB revealed an enhanced ability to induce MAP/ERK signaling while having a lower dependence on heparin when compared with FGF2-wt. Here we report the crystal structure of FGF2-STAB solved at 1.3 Å resolution. Protein stabilization is achieved by newly formed hydrophobic interactions, polar contacts, and one additional hydrogen bond. The overall structure of FGF2-STAB is similar to FGF2-wt and does not reveal information on the experimentally observed lower dependence on heparin. A noticeable difference in flexibility in the receptor binding region can explain the differences in signaling between FGF2-STAB and its wild-type counterpart. Our structural analysis provided molecular insights into the stabilization and unique biological properties of FGF2-STAB.

• Klíčová slova
• Protein flexibility, Stabilized fibroblast growth factor 2, X-ray structural analysis,
• Publikační typ
• časopisecké články MeSH

Glycosphingolipids (GSL) are a highly heterogeneous class of lipids representing the majority of the sphingolipid category. GSL are fundamental constituents of cellular membranes that have key roles in various biological processes, such as cellular signaling, recognition, and adhesion. Understanding the structural complexity of GSL is pivotal for unraveling their functional significance in a biological context, specifically their crucial role in the pathophysiology of various diseases. Mass spectrometry (MS) has emerged as a versatile and indispensable tool for the structural elucidation of GSL enabling a deeper understanding of their complex molecular structures and their key roles in cellular dynamics and patholophysiology. Here, we provide a thorough overview of MS techniques tailored for the analysis of GSL, emphasizing their utility in probing GSL intricate structures to advance our understanding of the functional relevance of GSL in health and disease. The application of tandem MS using diverse fragmentation techniques, including novel ion activation methodologies, in studying glycan sequences, linkage positions, and fatty acid composition is extensively discussed. Finally, we address current challenges, such as the detection of low-abundance species and the interpretation of complex spectra, and offer insights into potential solutions and future directions by improving MS instrumentation for enhanced sensitivity and resolution, developing novel ionization techniques, or integrating MS with other analytical approaches for comprehensive GSL characterization.

• Klíčová slova
• Derivatization, Fragmentation, Glycosphingolipids, Liquid chromatography, Mass spectrometry, Structural elucidation,
• MeSH
• glykosfingolipidy * chemie   analýza   MeSH
• hmotnostní spektrometrie * metody   MeSH
• lidé MeSH
• tandemová hmotnostní spektrometrie metody   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• glykosfingolipidy * MeSH

Glucans are most widespread polysaccharides in the nature. There is a large diversity in their molecular weight and configuration depending on the original source. According to the anomeric structure of glucose units it is possible to distinguish linear and branched α-, β- as well as mixed α,β-glucans with various glycoside bond positions and molecular masses. Isolation of glucans from raw sources needs removal of ballast compounds including proteins, lipids, polyphenols and other polysaccharides. Purity control of glucan fractions is necessary to evaluate the isolation and purification steps; more rigorous structural analyses of purified polysaccharides are required to clarify their structure. A set of spectroscopic, chemical and separation methods are used for this purpose. Among them, NMR spectroscopy is known as a powerful tool in structural analysis of glucans both in solution and in solid state. Along with chemolytic methods [methylation analysis (MA), periodate oxidation, partial chemical or enzymatic hydrolysis, etc.], correlation NMR experiments are able to determine the exact structure of tested polysaccharides. Vibration spectroscopic methods (FTIR, Raman) are sensitive to anomeric structure of glucans and can be used for purity control as well. Molecular weight distribution, homogeneity and branching of glucans can be estimated by size-exclusion chromatography (SEC), laser light scattering (LLS) and viscometry.

• Klíčová slova
• Glucans, NMR spectroscopy, chemolytic methods, molecular weight, purity, structure, vibration spectroscopy,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

A detailed description of the dnatco.datmos.org web server implementing the universal structural alphabet of nucleic acids is presented. It is capable of processing any mmCIF- or PDB-formatted files containing DNA or RNA molecules; these can either be uploaded by the user or supplied as the wwPDB or PDB-REDO structural database access code. The web server performs an assignment of the nucleic acid conformations and presents the results for the intuitive annotation, validation, modeling and refinement of nucleic acids.

• Klíčová slova
• annotation, nucleic acids, refinement, structural alphabets, validation,
• MeSH
• databáze nukleových kyselin MeSH
• DNA chemie   MeSH
• internet MeSH
• konformace nukleové kyseliny MeSH
• molekulární modely MeSH
• RNA chemie   MeSH
• software * MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA MeSH
• RNA MeSH

Aldehyde dehydrogenases (ALDHs) constitute a superfamily of NAD(P)+-dependent enzymes, which detoxify aldehydes produced in various metabolic pathways to the corresponding carboxylic acids. Among the 19 human ALDHs, the cytosolic ALDH9A1 has so far never been fully enzymatically characterized and its structure is still unknown. Here, we report complete molecular and kinetic properties of human ALDH9A1 as well as three crystal forms at 2.3, 2.9, and 2.5 Å resolution. We show that ALDH9A1 exhibits wide substrate specificity to aminoaldehydes, aliphatic and aromatic aldehydes with a clear preference for γ-trimethylaminobutyraldehyde (TMABAL). The structure of ALDH9A1 reveals that the enzyme assembles as a tetramer. Each ALDH monomer displays a typical ALDHs fold composed of an oligomerization domain, a coenzyme domain, a catalytic domain, and an inter-domain linker highly conserved in amino-acid sequence and folding. Nonetheless, structural comparison reveals a position and a fold of the inter-domain linker of ALDH9A1 never observed in any other ALDH so far. This unique difference is not compatible with the presence of a bound substrate and a large conformational rearrangement of the linker up to 30 Å has to occur to allow the access of the substrate channel. Moreover, the αβE region consisting of an α-helix and a β-strand of the coenzyme domain at the dimer interface are disordered, likely due to the loss of interactions with the inter-domain linker, which leads to incomplete β-nicotinamide adenine dinucleotide (NAD+) binding pocket.

• Klíčová slova
• 3-aminopropionaldehyde, 4-trimethylaminobutyraldehyde, Homo sapiens, X‐ray crystallography, aldehyde dehydrogenase, structure-function,
• MeSH
• aldehyddehydrogenasa antagonisté a inhibitory   chemie   genetika   ultrastruktura   MeSH
• katalytická doména genetika   MeSH
• kinetika MeSH
• konformace proteinů * MeSH
• krystalografie rentgenová MeSH
• lidé MeSH
• NAD genetika   MeSH
• sekundární struktura proteinů MeSH
• sekvence aminokyselin genetika   MeSH
• substrátová specifita genetika   MeSH
• vazebná místa genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aldehyddehydrogenasa MeSH
• ALDH9A1 protein, human MeSH Prohlížeč
• NAD MeSH

This article is devoted to the characterization of a new Co-W-Al alloy prepared by an aluminothermic reaction. This alloy is used for the subsequent preparation of a special composite nanopowder and for the surface coating of aluminum, magnesium, or iron alloys. Due to the very high temperature (2000 °C-3000 °C) required for the reaction, thermite was added to the mixture. Pulverized coal was also added in order to obtain the appropriate metal carbides (Co, W, Ti), which increase hardness, resistance to abrasion, and the corrosion of the coating and have good high temperature properties. The phase composition of the alloy prepared by the aluminothermic reaction showed mainly cobalt, tungsten, and aluminum, as well as small amounts of iron, titanium, and calcium. No carbon was identified using this method. The microstructure of this alloy is characterized by a cobalt matrix with smaller regular and irregular carbide particles doped by aluminum.

• Klíčová slova
• Co-W-Al alloy, EDS analysis, aluminothermic reaction, carbides, chemical composition, scanning electron microscopy, structural analysis,
• Publikační typ
• časopisecké články MeSH

Longitudinal comorbidity of depression and cognitive impairment has been reported by number of epidemiological studies but the underlying mechanisms explaining the link between affective problems and cognitive decline are not very well understood. Imaging studies have typically investigated patients with major depressive disorder (MDD) and mild cognitive impairment (MCI) separately and thus have not identified a structural brain signature common to these conditions that may illuminate potentially targetable shared biological mechanisms. We performed a meta-analysis of. 48 voxel-based morphometry (VBM) studies of individuals with MDD, MCI, and age-matched controls and demonstrated that MDD and MCI patients had shared volumetric reductions in a number of regions including the insula, superior temporal gyrus (STG), inferior frontal gyrus, amygdala, hippocampus, and thalamus. We suggest that the shared volumetric reductions in the insula and STG might reflect communication deficits and infrequent participation in mentally or socially stimulating activities, which have been described as risk factors for both MCI and MDD. We also suggest that the disease-specific structural changes might reflect the disease-specific symptoms such as poor integration of emotional information, feelings of helplessness and worthlessness, and anhedonia in MDD. These findings could contribute to better understanding of the origins of MDD-MCI comorbidity and facilitate development of early interventions.

• Klíčová slova
• Major depressive disorder, Meta-analysis, Mild cognitive impairment, Shared volumetric reductions, structural magnetic resonance imaging (MRI), voxel-based morphometry (VBM),
• MeSH
• deprese MeSH
• depresivní porucha unipolární * diagnostické zobrazování   MeSH
• kognitivní dysfunkce * diagnostické zobrazování   MeSH
• lidé MeSH
• magnetická rezonanční tomografie MeSH
• mozek diagnostické zobrazování   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• metaanalýza MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

BACKGROUND: Structural variants (SVs) represent an important source of genetic variation. One of the most critical problems in their detection is breakpoint uncertainty associated with the inability to determine their exact genomic position. Breakpoint uncertainty is a characteristic issue of structural variants detected via short-read sequencing methods and complicates subsequent population analyses. The commonly used heuristic strategy reduces this issue by clustering/merging nearby structural variants of the same type before the data from individual samples are merged. RESULTS: We compared the two most used dissimilarity measures for SV clustering in terms of Mendelian inheritance errors (MIE), kinship prediction, and deviation from Hardy-Weinberg equilibrium. We analyzed the occurrence of Mendelian-inconsistent SV clusters that can be collapsed into one Mendelian-consistent SV as a new measure of dataset consistency. We also developed a new method based on constrained clustering that explicitly identifies these types of clusters. CONCLUSIONS: We found that the dissimilarity measure based on the distance between SVs breakpoints produces slightly better results than the measure based on SVs overlap. This difference is evident in trivial and corrected clustering strategy, but not in constrained clustering strategy. However, constrained clustering strategy provided the best results in all aspects, regardless of the dissimilarity measure used.

• Klíčová slova
• Breakpoints uncertainty problem, Constrained clustering, Mendelian inheritance error, Structural variants, Whole genome sequencing,
• MeSH
• genom lidský * MeSH
• genomika MeSH
• lidé MeSH
• nejistota MeSH
• shluková analýza MeSH
• strukturální variace genomu * MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

We report the crystal structure of the SARS-CoV-2 putative primase composed of the nsp7 and nsp8 proteins. We observed a dimer of dimers (2:2 nsp7-nsp8) in the crystallographic asymmetric unit. The structure revealed a fold with a helical core of the heterotetramer formed by both nsp7 and nsp8 that is flanked with two symmetry-related nsp8 β-sheet subdomains. It was also revealed that two hydrophobic interfaces one of approx. 1340 Å2 connects the nsp7 to nsp8 and a second one of approx. 950 Å2 connects the dimers and form the observed heterotetramer. Interestingly, analysis of the surface electrostatic potential revealed a putative RNA binding site that is formed only within the heterotetramer.

• Klíčová slova
• Crystal structure, Primase, RNA, SARS-CoV-2,
• MeSH
• Betacoronavirus chemie   MeSH
• DNA-primasa chemie   metabolismus   MeSH
• konformace proteinů MeSH
• koronavirová RNA-replikasa MeSH
• krystalografie rentgenová MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• multiproteinové komplexy MeSH
• RNA metabolismus   MeSH
• SARS-CoV-2 MeSH
• vazebná místa MeSH
• virové nestrukturální proteiny chemie   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA-primasa MeSH
• koronavirová RNA-replikasa MeSH
• multiproteinové komplexy MeSH
• NS8 protein, SARS-CoV-2 MeSH Prohlížeč
• NSP7 protein, SARS-CoV-2 MeSH Prohlížeč
• RNA MeSH
• virové nestrukturální proteiny MeSH