GTP-cyclohydrolases of microorganisms and their role in metabolism

The properties, regularities of biosynthesis and biochemical functions are considered of GTP-cyclohydrolases of microorganisms. The existence of two groups of these enzymes is established. The first group enzymes convert GTP into 7,8-dihydroneopterin-triphosphate and formiate. They participate in biosynthesis of tetrahydrofolic acid, tetrahydrobiopterin, molybdenic cofactor, pyrrolopyrimidine antibiotics and in a series of pigments. Representatives of the second group of cyclohydrolases convert GTP into 2,5-diamino-4-oxy-6-ribosylaminopyrimidine-5'-phosphate, formiate and pyrophosphate. They catalyze the first stages of formation of 6,7-dimethyl-8-ribityllumazine, flavins and their derivatives, toxoflavin (azapteridine antibiotics). The regulation of biosynthesis and activity of GTP-cyclohydrolases is studied only for individual enzymes of this group.     

Overexpression of a novel member of the mitochondrial carrier family rescues defects in both DNA and RNA metabolism in yeast mitochondria

The PIF1 and MRS2 gene products have previously been shown to be essential for mitochondrial DNA maintenance at elevated temperatures and mitochondrial group II intron splicing, respectively, in the yeast Saccharomyces cerevisiae. A multicopy suppressor capable of rescuing the respiratory deficient phenotype associated with null alleles of either gene has been isolated. This suppressor is a nuclear gene that was called RIM2/MRS12. The RIM2/MRS12 gene encodes a predicted protein of 377 amino acids that is essential for mitochondrial DNA metabolism and proper cell growth. Inactivation of this gene causes the total loss of mitochondrial DNA and, compared to wild-type rho^o controls, a slow-growth phenotype on media containing glucose. Analysis of the RIM2/MRS12 protein sequence suggests that RIM2/MRS12 encodes a novel member of the mitochondrial carrier family. In particular, a typical triplicate structure, where each repeat consists of two putative transmembrane segments separated by a hydrophilic loop, can be deduced from amino acid sequence comparisons and the hydropathy profile of RIM2/MRS12. Antibodies directed against the aminoterminus of RIM2/MRS12 detect this protein in mitochondria. The function of the RIM2/MRS12 protein and the substrates it might transport are discussed.     

Bacterial haloperoxidases and their role in secondary metabolism

Bacteria produce a large number of different halogenated secondary metabolites. Haloperoxidases are believed to be the enzymes responsible for the halogenation reaction. Two classes of haloperoxidases, heme and nonheme, were isolated from different bacteria and their role in the biosynthesis of halogenated secondary metabolites was investigated. Two genes of bacterial haloperoxidases were cloned and can now be used to produce large quantities of the enzymes.     

The Shwachman-Bodian-Diamond syndrome protein family is involved in RNA metabolism

A combination of structural, biochemical, and genetic studies in model organisms was used to infer a cellular role for the human protein (SBDS) responsible for Shwachman-Bodian-Diamond syndrome. The crystal structure of the SBDS homologue in Archaeoglobus fulgidus, AF0491, revealed a three domain protein. The N-terminal domain, which harbors the majority of disease-linked mutations, has a novel three-dimensional fold. The central domain has the common winged helix-turn-helix motif, and the C-terminal domain shares structural homology with known RNA-binding domains. Proteomic analysis of the SBDS sequence homologue in Saccharomyces cerevisiae, YLR022C, revealed an association with over 20 proteins involved in ribosome biosynthesis. NMR structural genomics revealed another yeast protein, YHR087W, to be a structural homologue of the AF0491 N-terminal domain. Sequence analysis confirmed them as distant sequence homologues, therefore related by divergent evolution. Synthetic genetic array analysis of YHR087W revealed genetic interactions with proteins involved in RNA and rRNA processing including Mdm20/Nat3, Nsr1, and Npl3. Our observations, taken together with previous reports, support the conclusion that SBDS and its homologues play a role in RNA metabolism.     

Ribonucleotide reductase--transition enzymes from RNA metabolism to DNA metabolism]

Ribonucleotide reductases catalyze the reduction of ribonucleotides to deoxyribonucleotides, this reaction being vitally important for all organisms. The enzyme provides a link between RNA and DNA metabolisms. Several forms of the enzyme occur in nature and those differ in their structure and catalytic mechanisms. While the direct reduction of ribonucleotides via a radical mechanism is a general mode of dNTP synthesis in all organisms, the way in which this is achieved varies. The ability of the enzyme to control DNA synthesis is of considerable clinical interest. Selective inhibition of DNA synthesis is desired, for example, in clinical applications, such as restriction of tumour growth or virus replication.     

Methallothioneins and their role in the metabolism and toxicity of metals.

Recent investigations have provided considerable new information regarding the biological role of metallothioneins. The synthesis of this protein is induced in cells by certain metals. It can tightly bind with zinc, copper, cadmium, mercury or silver reducing the availability of diffusible forms of these metals within cells and therefore decreasing their toxic potential. The metallothioneins may also have an important role in regulating the normal absorption and homeostasis of zinc and copper. It is paradoxical, however, in that a protein synthesized within the cell to reduce toxicity, may, in itself, be toxic when excreted or leaked out from the cell to the extracellular space. Further studies are required to elucidate the mechanisms involved in these effects.     

The emerging role of RNA and DNA editing in cancer

Nucleotide sequence modification through single base editing in animals is emerging as an important player in tumorigenesis. RNA editing especially has increased greatly during mammalian evolution and modulates diverse cellular functions presumably in a context-dependent manner. Sequence editing impacts development, including pluripotency and hematopoiesis, and multiple recent studies have shown that dysregulation of editing is associated with tumor biology. Much is yet to be learned about the role of sequence editing in human biology but this process is a critical modulator of cell regulation and may present an attractive option for therapeutic intervention in cancer in the future.SignificanceSequence editing provides an additional regulatory layer of cancer initiation and progression that may be amenable to therapeutic design. Although editing of both RNA and DNA substrates has been known to occur for some time, the extent and implications of these modifications have been grossly underappreciated until recent genome-wide and disease-association studies were reported. This review highlights the cellular processes controlled by sequence editing, their implications in normal and cancerous states and considers potential targeted therapeutic strategies.     

Vacuolar transporters and their essential role in plant metabolism

Following the unequivocal demonstration that plants contain at least two types of vacuoles, scientists studying this organelle have realized that the plant 'vacuome' is far more complex than they expected. Some fully developed cells contain at least two large vacuoles, with different functions. Remarkably, even a single vacuole may be subdivided and fulfil several functions, which are supported in part by the vacuolar membrane transport systems. Recent studies, including proteomic analyses for several plant species, have revealed the tonoplast transporters and their involvement in the nitrogen storage, salinity tolerance, heavy metal homeostasis, calcium signalling, guard cell movements, and the cellular pH homeostasis. It is clear that vacuolar transporters are an integrated part of a complex cellular network that enables a plant to react properly to changing environmental conditions, to save nutrients and energy in times of plenty, and to maintain optimal metabolic conditions in the cytosol. An overview is given of the main features of the transporters present in the tonoplast of plant cells in terms of their function, regulation, and relationships with the microheterogeneity of the vacuome.     

Endogenous cannabinoids: metabolism and their role in reproduction

Over the past two decades a number of endogenous compounds that act as ligands for the cannabinoid receptors has been discovered. In analogy with the "endorphins" these compounds have been called "endocannabinoids". Endocannabinoids have been demonstrated in many mammalian tissues including humans and are widely distributed in the CNS, peripheral nerves, uterus, leukocytes, spleen and testicles. The uterus contains the highest levels of anandamide, the first discovered endocannabinoid, suggesting an important role for this substance in reproduction. Several studies have shown anandamide to be involved in the regulation of implantation and reduced activity of the enzyme that degrades anandamide has been associated with early pregnancy loss in humans. The bulk of the literature concerning endocannabinoids is based upon anandamide related studies; therefore, in this review we focus on the metabolism of anandamide and its role in reproduction.     

Ubiquitin-binding domains and their role in the DNA damage response

The modification of eukaryotic proteins by covalent attachment of ubiquitin is a versatile signaling event with a wide range of possible consequences. Canonical poly-ubiquitination by Lys-48 linked chains usually destines a protein for degradation by the proteasome. By contrast, attachment of a single ubiquitin or ubiquitin chains linked through Lys-63 or Lys-6 serves a non-proteolytic role. Over the last years, evidence has accumulated that several nuclear proteins become ubiquitinated in response to DNA damage. Typically, these proteins carry mono-ubiquitin or non-classical ubiquitin chains and are localized close to the site of DNA damage. Of particular interest are PCNA and the variant histone H2AX, two key proteins whose ubiquitination serves to recruit factors needed by the cell to cope with the damage. A prerequisite for docking effector proteins to the site of the lesion is the detection of a specific ubiquitin modification, a process that can be mediated by a range of dedicated ubiquitin-binding domains (UBDs). As the same types of ubiquitin modification are involved in entirely different processes, the recognition of the ubiquitin mark has to go along with the recognition of the modified protein. Thus, ubiquitin-binding domains gain their specificity through combination with other recognition domains and motifs. This review discusses ubiquitin-binding domains relevant to the DNA damage response, including their binding mode, their specificity, and their interdependence with other factors. For several repair pathways, current knowledge of the events downstream of the ubiquitin mark is sketchy. A closer look at orphan UBD proteins might lead to the identification of missing pieces in the DNA response puzzle.     

Neuroglobin and cytoglobin in search of their role in the vertebrate globin family

Neuroglobin and cytoglobin are two recent additions to the family of heme-containing respiratory proteins of man and other vertebrates. Here, we review the present state of knowledge of the structures, ligand binding kinetics, evolution and expression patterns of these two proteins. These data provide a first glimpse into the possible physiological roles of these globins in the animal's metabolism. Both, neuroglobin and cytoglobin are structurally similar to myoglobin, although they contain distinct cavities that may be instrumental in ligand binding. Kinetic and structural studies show that neuroglobin and cytoglobin belong to the class of hexa-coordinated globins with a biphasic ligand-binding kinetics. Nevertheless, their oxygen affinities resemble that of myoglobin. While neuroglobin is evolutionarily related to the invertebrate nerve-globins, cytoglobin shares a more recent common ancestry with myoglobin. Neuroglobin expression is confined mainly to brain and a few other tissues, with the highest expression observed in the retina. Present evidence points to an important role of neuroglobin in neuronal oxygen homeostasis and hypoxia protection, though other functions are still conceivable. Cytoglobin is predominantly expressed in fibroblasts and related cell types, but also in distinct nerve cell populations. Much less is known about its function, although in fibroblasts it might be involved in collagen synthesis.     

Human carboxylesterases and their role in xenobiotic and endobiotic metabolism

Carboxylesterases (CEs) are traditionally regarded as xenobiotic metabolizing enzymes that hydrolyze esterified xenobiotics to alcohol and carboxylic acid products. However, there is a growing appreciation for the role of CEs in the processing of endobiotics, including cholesteryl esters and triacylglycerols. Human liver microsomes (HLMs) are often used in reaction phenotyping studies to discern interindividual variability in xenobiotic metabolism. The two major CE isoforms expressed in human liver are hCE1 and hCE2. These two isoforms are different gene products. We have begun studies to evaluate the CE phenotype' of human liver samples, i.e. to determine both the levels of hCE1 and hCE2 protein and the hydrolytic activity of each. We have previously shown that there is little variation in hCE1 protein expression in HLM samples from 11 individuals [a 1.3-fold difference between the highest and lowest individuals; coefficient of variation (CV), 9%]. hCE2 protein expression in individual HLMs is only slightly more variable than hCE1 (2.3-fold difference between the highest and lowest individuals; CV, 36%). However, hCE1 protein is found in 46-fold higher amounts in HLMs than hCE2 protein (64.4 +/- 16.5 microg hCE1/mg microsomal protein compared to 1.4 +/- 0.2 microg hCE2/mg microsomal protein). The hydrolytic activity specifically attributable to hCE1 and hCE2 in individual HLMs was measured using bioresmethrin (a pyrethroid insecticide hydrolyzed specifically by hCE1, but not by hCE2) and procaine (an analgesic drug hydrolyzed by hCE2, but not by hCE1). The hydrolytic activity of individual HLMs toward bioresmethrin and procaine did not correlate with the protein content of hCE1 and hCE2. Thus, the mere abundance of CE proteins is not a good predictor of CE activity in HLMs. Identification of the factors that lead to altered CE activities in HLMs will be important to characterize since several pharmaceutical agents, environmental toxicants, and endobiotics are metabolized by these enzymes.