Evolutionary and molecular basis of ADP-ribosylation reversal by zinc-dependent macrodomains.

Dynamic ADP-ribosylation signaling is a crucial pathway that controls fundamental cellular processes, in particular, the response to cellular stresses such as DNA damage, reactive oxygen species, and infection. In some pathogenic microbes, the response to oxidative stress is controlled by a SirTM/zinc-containing macrodomain (Zn-Macro) pair responsible for establishment and removal of the modification, respectively. Targeting this defence mechanism against the host's innate immune response may lead to novel approaches to support the fight against emerging antimicrobial resistance.

The Crucial Role of Breeder and Dog Owners Associations in Safeguarding Genetic Heritage of Endangered Balearic Dog Breeds: Gender Preference and Registry Adscription.

This study delves into the complex relationships between indigenous dog breeds in the Balearic Isles and their human counterparts, specifically breeders and owners. Using Canonical Correlation Analysis, the research examines variables such as breed registries and the number of breeders/owners, uncovering significant correlations within registries. For example, an increase in female auxiliary registrations corresponds to a decline in foundational registrations, indicating shifts in breed documentation dynamics.

Molecular basis for the reversible ADP-ribosylation of guanosine bases.

Modification of nucleic acids by ADP-ribosylation is catalyzed by various ADP-ribosyltransferases, including the DarT enzyme. The latter is part of the bacterial toxin-antitoxin (TA) system DarTG, which was shown to provide control of DNA replication and bacterial growth as well as protection against bacteriophages. Two subfamilies have been identified, DarTG1 and DarTG2, which are distinguished by their associated antitoxins.

Serine ADP-ribosylation in Drosophila provides insights into the evolution of reversible ADP-ribosylation signalling.

In the mammalian DNA damage response, ADP-ribosylation signalling is of crucial importance to mark sites of DNA damage as well as recruit and regulate repairs factors. Specifically, the PARP1:HPF1 complex recognises damaged DNA and catalyses the formation of serine-linked ADP-ribosylation marks (mono-Ser-ADPr), which are extended into ADP-ribose polymers (poly-Ser-ADPr) by PARP1 alone. Poly-Ser-ADPr is reversed by PARG, while the terminal mono-Ser-ADPr is removed by ARH3.

ERCC6L2 mitigates replication stress and promotes centromere stability.

Structurally complex genomic regions, such as centromeres, are inherently difficult to duplicate. The mechanism behind centromere inheritance is not well understood, and one of the key questions relates to the reassembly of centromeric chromatin following DNA replication. Here, we define ERCC6L2 as a key regulator of this process.

Molecular basis for DarT ADP-ribosylation of a DNA base.

ADP-ribosyltransferases use NAD to catalyse substrate ADP-ribosylation, and thereby regulate cellular pathways or contribute to toxin-mediated pathogenicity of bacteria. Reversible ADP-ribosylation has traditionally been considered a protein-specific modification, but recent in vitro studies have suggested nucleic acids as targets. Here we present evidence that specific, reversible ADP-ribosylation of DNA on thymidine bases occurs in cellulo through the DarT-DarG toxin-antitoxin system, which is found in a variety of bacteria (including global pathogens such as Mycobacterium tuberculosis, enteropathogenic Escherichia coli and Pseudomonas aeruginosa).

Mechanistic insights into the three steps of poly(ADP-ribosylation) reversal.

Poly(ADP-ribosyl)ation (PAR) is a versatile and complex posttranslational modification composed of repeating units of ADP-ribose arranged into linear or branched polymers. This scaffold is linked to the regulation of many of cellular processes including the DNA damage response, alteration of chromatin structure and Wnt signalling. Despite decades of research, the principles and mechanisms underlying all steps of PAR removal remain actively studied.

HPF1 completes the PARP active site for DNA damage-induced ADP-ribosylation.

The anti-cancer drug target poly(ADP-ribose) polymerase 1 (PARP1) and its close homologue, PARP2, are early responders to DNA damage in human cells. After binding to genomic lesions, these enzymes use NAD to modify numerous proteins with mono- and poly(ADP-ribose) signals that are important for the subsequent decompaction of chromatin and the recruitment of repair factors. These post-translational modifications are predominantly serine-linked and require the accessory factor HPF1, which is specific for the DNA damage response and switches the amino acid specificity of PARP1 and PARP2 from aspartate or glutamate to serine residues.

Structural and Functional Characterization of Three Novel Fungal Amylases with Enhanced Stability and pH Tolerance.

Amylases are probably the best studied glycoside hydrolases and have a huge biotechnological value for industrial processes on starch. Multiple amylases from fungi and microbes are currently in use. Whereas bacterial amylases are well suited for many industrial processes due to their high stability, fungal amylases are recognized as safe and are preferred in the food industry, although they lack the pH tolerance and stability of their bacterial counterparts.

Structural insight into industrially relevant glucoamylases: flexible positions of starch-binding domains.

Glucoamylases are one of the most important classes of enzymes in the industrial degradation of starch biomass. They consist of a catalytic domain and a carbohydrate-binding domain (CBM), with the latter being important for the interaction with the polymeric substrate. Whereas the catalytic mechanisms and structures of the individual domains are well known, the spatial arrangement of the domains with respect to each other and its influence on activity are not fully understood.

Structural insights into the function of ZRANB3 in replication stress response.

Strategies to resolve replication blocks are critical for the maintenance of genome stability. Among the factors implicated in the replication stress response is the ATP-dependent endonuclease ZRANB3. Here, we present the structure of the ZRANB3 HNH (His-Asn-His) endonuclease domain and provide a detailed analysis of its activity.

The Toxin-Antitoxin System DarTG Catalyzes Reversible ADP-Ribosylation of DNA.

The discovery and study of toxin-antitoxin (TA) systems helps us advance our understanding of the strategies prokaryotes employ to regulate cellular processes related to the general stress response, such as defense against phages, growth control, biofilm formation, persistence, and programmed cell death. Here we identify and characterize a TA system found in various bacteria, including the global pathogen Mycobacterium tuberculosis. The toxin of the system (DarT) is a domain of unknown function (DUF) 4433, and the antitoxin (DarG) a macrodomain protein.

The role of ADP-ribosylation in regulating DNA interstrand crosslink repair.

ADP-ribosylation by ADP-ribosyltransferases (ARTs) has a well-established role in DNA strand break repair by promoting enrichment of repair factors at damage sites through ADP-ribose interaction domains. Here, we exploit the simple eukaryote Dictyostelium to uncover a role for ADP-ribosylation in regulating DNA interstrand crosslink repair and redundancy of this pathway with non-homologous end-joining (NHEJ). In silico searches were used to identify a protein that contains a permutated macrodomain (which we call aprataxin/APLF-and-PNKP-like protein; APL).

Three-dimensional structures of two heavily N-glycosylated Aspergillus sp. family GH3 β-D-glucosidases.

The industrial conversion of cellulosic plant biomass into useful products such as biofuels is a major societal goal. These technologies harness diverse plant degrading enzymes, classical exo- and endo-acting cellulases and, increasingly, cellulose-active lytic polysaccharide monooxygenases, to deconstruct the recalcitrant β-D-linked polysaccharide. A major drawback with this process is that the exo-acting cellobiohydrolases suffer from severe inhibition from their cellobiose product.

The crystal structure of the Hazara virus nucleocapsid protein.

Background: Hazara virus (HAZV) is a member of the Bunyaviridae family of segmented negative stranded RNA viruses, and shares the same serogroup as Crimean-Congo haemorrhagic fever virus (CCHFV). CCHFV is responsible for fatal human disease with a mortality rate approaching 30 %, which has an increased recent incidence within southern Europe. There are no preventative or therapeutic treatments for CCHFV-mediated disease, and thus CCHFV is classified as a hazard group 4 pathogen.

Identification of a Class of Protein ADP-Ribosylating Sirtuins in Microbial Pathogens.

Sirtuins are an ancient family of NAD(+)-dependent deacylases connected with the regulation of fundamental cellular processes including metabolic homeostasis and genome integrity. We show the existence of a hitherto unrecognized class of sirtuins, found predominantly in microbial pathogens. In contrast to earlier described classes, these sirtuins exhibit robust protein ADP-ribosylation activity.

Synthesis of dimeric ADP-ribose and its structure with human poly(ADP-ribose) glycohydrolase.

Poly(ADP-ribosyl)ation is a common post-translational modification that mediates a wide variety of cellular processes including DNA damage repair, chromatin regulation, transcription, and apoptosis. The difficulty associated with accessing poly(ADP-ribose) (PAR) in a homogeneous form has been an impediment to understanding the interactions of PAR with poly(ADP-ribose) glycohydrolase (PARG) and other binding proteins. Here we describe the chemical synthesis of the ADP-ribose dimer, and we use this compound to obtain the first human PARG substrate-enzyme cocrystal structure.

Probing bunyavirus N protein oligomerisation using mass spectrometry.

Rationale: Bunyaviruses have become a major threat to both humans and livestock in Europe and the Americas. The nucleocapsid (N) protein of these viruses is key to the replication cycle and knowledge of the N oligomerisation state is central to understanding the viral lifecycle and for development of therapeutic strategies.

Methods: Bunyamwera virus and Schmallenberg virus N proteins (BUNV-N and SBV-N) were expressed recombinantly in E.

Crystal structure of the essential transcription antiterminator M2-1 protein of human respiratory syncytial virus and implications of its phosphorylation.

The M2-1 protein of the important pathogen human respiratory syncytial virus is a zinc-binding transcription antiterminator that is essential for viral gene expression. We present the crystal structure of full-length M2-1 protein in its native tetrameric form at a resolution of 2.5 Å.

Mechanism of protein kinetic stabilization by engineered disulfide crosslinks.

The impact of disulfide bonds on protein stability goes beyond simple equilibrium thermodynamics effects associated with the conformational entropy of the unfolded state. Indeed, disulfide crosslinks may play a role in the prevention of dysfunctional association and strongly affect the rates of irreversible enzyme inactivation, highly relevant in biotechnological applications. While these kinetic-stability effects remain poorly understood, by analogy with proposed mechanisms for processes of protein aggregation and fibrillogenesis, we propose that they may be determined by the properties of sparsely-populated, partially-unfolded intermediates.

Degradation of phytate by the 6-phytase from Hafnia alvei: a combined structural and solution study.

Phytases hydrolyse phytate (myo-inositol hexakisphosphate), the principal form of phosphate stored in plant seeds to produce phosphate and lower phosphorylated myo-inositols. They are used extensively in the feed industry, and have been characterised biochemically and structurally with a number of structures in the PDB. They are divided into four distinct families: histidine acid phosphatases (HAP), β-propeller phytases, cysteine phosphatases and purple acid phosphatases and also split into three enzyme classes, the 3-, 5- and 6-phytases, depending on the position of the first phosphate in the inositol ring to be removed.

Nucleocapsid protein structures from orthobunyaviruses reveal insight into ribonucleoprotein architecture and RNA polymerization.

All orthobunyaviruses possess three genome segments of single-stranded negative sense RNA that are encapsidated with the virus-encoded nucleocapsid (N) protein to form a ribonucleoprotein (RNP) complex, which is uncharacterized at high resolution. We report the crystal structure of both the Bunyamwera virus (BUNV) N-RNA complex and the unbound Schmallenberg virus (SBV) N protein, at resolutions of 3.20 and 2.

Structure, function, and evolution of the Crimean-Congo hemorrhagic fever virus nucleocapsid protein.

Crimean-Congo hemorrhagic fever virus (CCHFV) is an emerging tick-borne virus of the Bunyaviridae family that is responsible for a fatal human disease for which preventative or therapeutic measures do not exist. We solved the crystal structure of the CCHFV strain Baghdad-12 nucleocapsid protein (N), a potential therapeutic target, at a resolution of 2.1 Å.

Structure and activity of Paenibacillus polymyxa xyloglucanase from glycoside hydrolase family 44.

The enzymatic degradation of plant polysaccharides is emerging as one of the key environmental goals of the early 21st century, impacting on many processes in the textile and detergent industries as well as biomass conversion to biofuels. One of the well known problems with the use of nonstarch (nonfood)-based substrates such as the plant cell wall is that the cellulose fibers are embedded in a network of diverse polysaccharides, including xyloglucan, that renders access difficult. There is therefore increasing interest in the "accessory enzymes," including xyloglucanases, that may aid biomass degradation through removal of "hemicellulose" polysaccharides.