DING Proteins Extend to the Extremophilic World.

The DING proteins are ubiquitous in the three domains of life, from mesophiles to thermo- and hyperthermophiles. They belong to a family of more than sixty members and have a characteristic N-terminus, DINGGG, which is considered a "signature" of these proteins. Structurally, they share a highly conserved phosphate binding site, and a three dimensional organization resembling the "Venus Flytrap", both reminding the ones of PstS proteins.

In , the Poly(ADP-Ribose) Polymerase-Like Thermoprotein Is a Multifunctional Enzyme.

In , Sso, the ADP-ribosylating thermozyme is known to carry both auto- and heteromodification of target proteins via short chains of ADP-ribose. Here, we provide evidence that this thermoprotein is a multifunctional enzyme, also showing ATPase activity. Electrophoretic and kinetic analyses were performed using NAD and ATP as substrates.

Automodified Poly(ADP-Ribose) Polymerase Analysisto Monitor DNA Damagein Peripheral Lymphocytes of Floriculturists Occupationally Exposed to Pesticides.

Increased DNA damage and the propension to cancer development, depend on the modulation of the mechanisms to control and maintain genomic integrity. Poly(ADP-Ribose)Polymerase activation and automodification are early responses to genotoxic stress. Upon binding to DNA strand breaks, the enzyme, a molecular DNA nick sensor, is hyperactivated: this is the first step in a series of events leading to either DNA repair or apoptosis.

Diastereomeric Recognition of 5',8-cyclo-2'-Deoxyadenosine Lesions by Human Poly(ADP-ribose) Polymerase 1 in a Biomimetic Model.

5',8-Cyclo-2'-deoxyadenosine (cdA), in the 5' and 5'diastereomeric forms, are typical non strand-break oxidative DNA lesions, induced by hydroxyl radicals, with emerging importance as a molecular marker. These lesions are exclusively repaired by the nucleotide excision repair (NER) mechanism with a low efficiency, thus readily accumulating in the genome. Poly(ADP-ribose) polymerase1 (PARP1) acts as an early responder to DNA damage and plays a key role as a nick sensor in the maintenance of the integrity of the genome by recognizing nicked DNA.

Comparison of the DING protein from the archaeon Sulfolobus solfataricus with human phosphate-binding protein and Pseudomonas fluorescence DING counterparts.

DING proteins represent a new group of 40 kDa-related members, ubiquitous in living organisms. The family also include the DING protein from Sulfolobus solfataricus, functionally related to poly(ADP-ribose) polymerases. Here, the archaeal protein has been compared with the human Phosphate-Binding Protein and the Pseudomonas fluorescence DING enzyme, by enzyme assays and immune cross-reactivity.

Hexadecenoic Fatty Acid Isomers in Human Blood Lipids and Their Relevance for the Interpretation of Lipidomic Profiles.

Monounsaturated fatty acids (MUFA) are emerging health biomarkers, and in particular the ratio between palmitoleic acid (9cis-16:1) and palmitic acid (16:0) affords the delta-9 desaturase index that is increased in obesity. Recently, other positional and geometrical MUFA isomers belonging to the hexadecenoic family (C16 MUFA) were found in circulating lipids, such as sapienic acid (6cis-16:1), palmitelaidic acid (9trans-16:1) and 6trans-16:1. In this work we report: i) the identification of sapienic acid as component of human erythrocyte membrane phospholipids with significant increase in morbidly obese patients (n = 50) compared with age-matched lean controls (n = 50); and ii) the first comparison of erythrocyte membrane phospholipids (PL) and plasma cholesteryl esters (CE) in morbidly obese patients highlighting that some of their fatty acid levels have opposite trends: increases of both palmitic and sapienic acids with the decrease of linoleic acid (9cis,12cis-18:2, omega-6) in red blood cell (RBC) membrane PL were reversed in plasma CE, whereas the increase of palmitoleic acid was similar in both lipid species.

A new facet of ADP-ribosylation reactions: SIRTs and PARPs interplay.

Nicotinamide Adenine Dinucleotide (NAD⁺) is known mainly as coenzyme of redox reactions for energy transduction and is consumed as substrate in regulatory reactions removing nicotinamide and producing ADP-ribose. Several families of ADP-ribose synthesizing enzymes use NAD⁺ as substrate and control processes like DNA repair, replication and transcription, chromatin structure, the activity of G-proteins and others. Since NAD⁺-dependent reactions involve degradation of the dinucleotide, a constant supply of the pyridinic substrate is required for its homeostasis.

Past and current topics on ADP-ribosylation reactions.

The milestone of Adenosine Diphosphate-ribosylation studies was the paper by Paul Mandel's group in 1960s, first describing a "sort" of polyadenylic acid synthesized upon addition of nicotinamide mononucleotide in rat liver nuclear extracts. Nicotinic Acid or Niacin is the precursor of Nicotinamide Adenin Dinucleotide. In 1960s this compound was known mainly as coenzyme of most redox processes in metabolism.

Vault-poly-ADP-ribose polymerase in the Octopus vulgaris brain: a regulatory factor of actin polymerization dynamic.

Our previous behavioural, biochemical and immunohistochemical analyses conducted in selected regions (supra/sub oesophageal masses) of the Octopus vulgaris brain detected a cytoplasmic poly-ADP-ribose polymerase (more than 90% of total enzyme activity). The protein was identified as the vault-free form of vault-poly-ADP-ribose polymerase. The present research extends and integrates the biochemical characterization of poly-ADP-ribosylation system, namely, reaction product, i.

Purification of the poly-ADP-ribose polymerase-like thermozyme from the archaeon Sulfolobus solfataricus.

Several different protocols have been developed to purify the ADP-ribosylating enzyme from Sulfolobus solfataricus. A number of techniques have been applied in regard to the crude homogenate preparation and protein extraction. Either mechanical cell lysis with DNAase digestion or freeze-thawing with sonication allowed to obtain fairly similar amounts of the thermozyme in the homogenate.

Physical exercise activates the poly(ADP-ribosyl)ation system in rat testes.

Given the high sensitivity of the male reproductive system to oxidative stress and to temperature changes, the amount of germ cell apoptosis and the activation of the poly(ADP-ribosyl)ation system (a very sensitive index of genotoxic stress) were evaluated in the testicular tissue of adult rats which underwent a 10-wk treadmill training, according to either a mild or a strong protocol; rats were sacrificed 24 h after the last training session or after a single bout of an additional stressing exercise (30 min of swimming). Controls were untrained rats (one resting group and one group with acute exercise). Both training and acute exercise increased marginally germ cell apoptotic indexes (caspase-induced poly(ADP-ribose) polymerase fragmentation and TUNEL-positive cells), while the activity of poly(ADP-ribose) polymerase and poly(ADP-ribose) glycohydrolase enzymes was affected in a way that suggests that acute exercise is associated with reversible genotoxic stress, and that training induces adaptive responses, as demonstrated by the activation of poly(ADP-ribose) polymerase system without subsequent increase in apoptosis.

Detecting clinical activity in systemic lupus erythematosus with an archaeal poly(ADP-ribose) polymerase-like thermozyme: a pivotal study.

The clinical usefulness of an immunotest was evaluated by using purified poly(adenosine diphosphate (ADP)-ribose) polymerase from Sulfolobus solfataricus (PARPSso) as an antigen to detect the presence of abnormal anti-PARP antibodies in the sera of patients with systemic lupus erythematosus (SLE) at different clinical stages. Sera from 44 patients with SLE, subgrouped on the basis of disease activity (16 with inactive disease, 28 with active disease) were analysed with a new immunotest to detect anti-PARP antibodies, and with an immunofluorescent (IIF) assay for antinuclear antibodies (ANA) detection. ANA detection by IIF revealed that sera of healthy subjects were negative, whereas sera from patients with SLE were positive in all cases (13 positive at 1:80, 15 at 1:160, 15 at 1:320, 1 at 1:640, v/v).

The ADP-ribosylation of Sulfolobus solfataricus Sso7 modulates protein/DNA interactions in vitro.

The 7 kDa Sso7 is a basic protein particularly abundant in Sulfolobus solfataricus and is involved in DNA assembly. This protein undergoes in vitro ADP-ribosylation by an endogenous poly(ADP-ribose) polymerase-like enzyme. The circular dichroism spectrum of purified ADP-ribosylated Sso7 shows that this modification stabilizes the prevalent protein beta-conformation, as suggested by shifting of negative ellipticity minimum to 220 nm.

Poly(ADP-ribosyl)ation of proteins and germ cell development in hyperthyroid rat testes.

The effect of increased serum levels of thyroid hormone (triiodothyronine, T3) on young rat testis spermatogenesis was studied by analysing molecular and morphological parameters. Hyperthyroidism was induced by either T3-treatment or 2- and 10-day cold exposure. The poly(ADP-ribosyl)ation of proteins catalysed by poly(ADP-ribose) polymerase, which is particularly active at specific stages of rat spermatogenesis, was analysed as molecular index of DNA damage and cell stress.

The ADP-ribosylating thermozyme from Sulfolobus solfataricus is a DING protein.

The partial amino acid sequence of the sulfolobal thermoprotein biochemically characterized as poly(ADP-ribose)polymerase-like enzyme overlaps those of DING proteins. This group of proteins, widely occurring in animals, plants and eubacteria, shows a characteristic and highly conserved N-terminus, DINGGGATL. The sequence of the N-terminal region and of the analyzed tryptic peptides of the sulfolobal thermozyme shows a high similarity with most of the DING proteins from databases.

NEMO-binding domain peptide inhibits proliferation of human melanoma cells.

Melanoma is the most aggressive form of skin cancer, it originates from melanocytes and its incidence has increased in the last decade. Recent advances in the understanding of the underlying biology of the progression of melanoma have identified key signalling pathways that are important in promoting melanoma tumourigenesis, thus providing dynamic targets for therapy. One such important target identified in melanoma tumour progression is the Nuclear Factor-kappaB (NF-kappaB) pathway.

The reaction of hydrogen atoms with methionine residues: A model of reductive radical stress causing tandem protein-lipid damage.

The occurrence of tandem damage, due to reductive radical stress involving proteins and lipids, is shown by using a biomimetic model. It is made of unsaturated lipid vesicle suspensions in phosphate buffer in the presence of methionine, either as a single amino acid or as part of a protein such as RNase A, which contains four methionine residues. The radical process starts with the formation of H(.

The (ADP-ribosyl)ation reaction in thermophilic bacteria.

Species of Alicyclobacillus, Bacillus and Thermus genera were selected in order to study the possible presence of the (ADP-ribosyl)ation system. These bacteria are thermophilic, aerobic, and were isolated from different geothermal sources. Both activity and expression of (ADP-ribosyl)ating proteins were tested in cells at different growth phases, and evidence of an active system was obtained in all analyzed microorganisms, with comparable enzymatic levels.

Investigation of radical-based damage of RNase A in aqueous solution and lipid vesicles.

The gamma-irradiation of bovine pancreatic ribonuclease A (RNase A) in aqueous solution were investigated at different doses by vibrational spectroscopy as well as enzymatic assay, electrophoresis, and HPLC analysis. Both functional and structural changes of the protein were caused by attack of H(*) atoms and (*)OH radicals. In particular, Raman spectroscopy was shown to be a useful tool in identifying conformational changes of the protein structure and amino acidic residues that are preferential sites of the radical attack (i.

Chromatin architecture and functions: the role(s) of poly(ADP-RIBOSE) polymerase and poly(ADPribosyl)ation of nuclear proteins.

Epigenetic states that allow chromatin fidelity inheritance can be mediated by several factors. One of them, histone variants and their modifications (including acetylation, methylation, phosphorylation, poly(ADP-ribosyl)ation, and ubiquitylation) create distinct patterns of signals read by other proteins, and are strictly related to chromatin remodelling, which is necessary for the specific expression of a gene, and for DNA repair, recombination, and replication. In the framework of chromatin-controlling factors, the poly(ADP-ribosyl)ation of nuclear proteins, catalysed by poly(ADP-ribose)polymerases (PARPs), has been implicated in the regulation of both physiological and pathological events (gene expression/amplification, cellular division/differentiation, DNA replication, malignant transformation, and apoptotic cell death).

Yeast (ADPribosyl)ation: revisiting a controversial question.

The controversy about the occurrence of an (ADPribosyl)ating activity in yeast is still standing up. Here we discuss this topic on the basis of results obtained with classic experiments proposed over years as basis to characterize an (ADPribosyl)ation system in any organism. Independent results obtained in two different laboratories were in line with each other and went towards the occurrence of an active (ADPribosyl)ating system in Saccharomyces cerevisiae.

Arachidonate geometrical isomers generated by thiyl radicals: the relationship with trans lipids detected in biological samples.

The presence of trans fatty acids in mammalians is attributed to exogenous sources; nevertheless, trans isomers could be easily formed by free radical-catalyzed isomerization processes in vivo. The isomerization of methyl arachidonate (all-cis isomer) catalyzed by thiyl radical is proposed as a methodology applicable in biochemical laboratories, which produces mono- and di-trans isomers. Carbon-13 nuclear magnetic resonance spectroscopy shows that the carbon atom in position 15 is characteristic for each mono- and di-trans isomer.

Histone H1-like protein and a testis-specific variant in the reproductive tracts of Octopus vulgaris.

In this study, we have identified a 28-kDa protein resembling the linker H1 in the testis and prostate of the reproductive system of Octopus vulgaris. This protein, OvH1, was partially purified by reverse phase high-pressure liquid chromatography (HPLC) of the perchloric acid extract from testis nuclei. It showed electrophoretic mobility, CD spectrum and amino acid composition highly comparable with those of the mammalian histone.

High stability binding of poly(ADPribose) polymerase-like thermozyme from S. solfataricus with circular DNA.

The poly(ADPribose) polymerase-like thermozyme from the hyperthermophilic archaeon S. solfataricus was found to bind DNA with high affinity and non-specifically. Binding was independent of base composition and length of the nucleic acid, and the protein showed a slight preference for the circular structure.