Oxygen-dependent Reactions Research Articles

Pulsed Multifrequency Electron Paramagnetic Resonance Spectroscopy Reveals Key Branch Points for One- vs Two-Electron Reactivity in Mn/Fe Proteins.

Traditionally, the ferritin-like superfamily of proteins was thought to exclusively use a diiron active site in catalyzing a diverse array of oxygen-dependent reactions. In recent years, novel redox-active cofactors featuring heterobimetallic Mn/Fe active sites have been discovered in both the radical-generating R2 subunit of class Ic (R2c) ribonucleotide reductases (RNRs) and the related R2-like ligand-binding oxidases (R2lox). However, the protein-specific factors that differentiate the radical reactivity of R2c from the C-H activation reactions of R2lox remain unknown. In this work, multifrequency pulsed electron paramagnetic resonance (EPR) spectroscopy and ligand hyperfine techniques in conjunction with broken-symmetry density functional theory calculations are used to characterize the molecular and electronic structures of two EPR-active intermediates trapped during aerobic assembly of the R2lox Mn/Fe cofactor. A MnIII(μ-O)(μ-OH)FeIII species is identified as the first EPR-active species and represents a common state between the two classes of redox-active Mn/Fe proteins. The species downstream from the MnIII(μ-O)(μ-OH)FeIII state exhibits unique EPR properties, including unprecedented spectral breadth and isotope-dependent g-tensors, which are attributed to a weakly coupled, hydrogen-bonded MnIII(μ-OH)FeIII species. This final intermediate precedes formation of the MnIII/FeIII resting state and is suggested to be relevant to understanding the endogenous reactivity of R2lox.

The role of hypoxia-inducible factors in the development of chronic pathology

This review highlights the current understanding of hypoxia-inducible factors (HIFs) role as regulators of oxygen-dependent reactions and inducers of genes expression in human organism. The focus is on the most significant relationships between the activation or inhibition of the HIFs intracellular system and development of the inflammatory process in various organs, chronic diseases of gastrointestinal tract, osteoarticular system, kidneys as well as hematological, endocrine and metabolic disorders.

Endogenous generation of nitro-fatty acid hybrids having dual nitrate ester (RONO2) and nitroalkene (RNO2) substituents

Organic nitrate esters, long-recognized therapies for cardiovascular disorders, have not been detected biologically. We characterize in rat stomach unsaturated fatty acid nitration reactions that proceed by generation of nitro-nitrate intermediates (NO2–ONO2-FA) via oxygen and nitrite dependent reactions. NO2–ONO2-lipids represent ∼70% of all nitrated lipids in the stomach and they decay in vitro at neutral or basic pH by the loss of the nitrate ester group (-ONO2) from the carbon backbone upon deprotonation of the α-carbon (pKa ∼7), yielding nitrate, nitrite, nitrosative species, and an electrophilic fatty acid nitroalkene product (NO2-FA). Of note, NO2-FA are anti-inflammatory and tissue-protective signaling mediators, which are undergoing Phase II trials for the treatment of kidney and pulmonary diseases. The decay of NO2–ONO2-FA occurs during intestinal transit and absorption, leading to the formation of NO2-FA that were subsequently detected in circulating plasma triglycerides. These observations provide new insight into unsaturated fatty acid nitration mechanisms, identify nitro-nitrate ester-containing lipids as intermediates in the formation of both secondary nitrogen oxides and electrophilic fatty acid nitroalkenes, and expand the scope of endogenous products stemming from metabolic reactions of nitrogen oxides.

De novo protein design, a retrospective.

Proteins are molecular machines whose function depends on their ability to achieve complex folds with precisely defined structural and dynamic properties. The rational design of proteins from first-principles, or de novo, was once considered to be impossible, but today proteins with a variety of folds and functions have been realized. We review the evolution of the field from its earliest days, placing particular emphasis on how this endeavor has illuminated our understanding of the principles underlying the folding and function of natural proteins, and is informing the design of macromolecules with unprecedented structures and properties. An initial set of milestones in de novo protein design focused on the construction of sequences that folded in water and membranes to adopt folded conformations. The first proteins were designed from first-principles using very simple physical models. As computers became more powerful, the use of the rotamer approximation allowed one to discover amino acid sequences that stabilize the desired fold. As the crystallographic database of protein structures expanded in subsequent years, it became possible to construct proteins by assembling short backbone fragments that frequently recur in Nature. The second set of milestones in de novo design involves the discovery of complex functions. Proteins have been designed to bind a variety of metals, porphyrins, and other cofactors. The design of proteins that catalyze hydrolysis and oxygen-dependent reactions has progressed significantly. However, de novo design of catalysts for energetically demanding reactions, or even proteins that bind with high affinity and specificity to highly functionalized complex polar molecules remains an importnant challenge that is now being achieved. Finally, the protein design contributed significantly to our understanding of membrane protein folding and transport of ions across membranes. The area of membrane protein design, or more generally of biomimetic polymers that function in mixed or non-aqueous environments, is now becoming increasingly possible.

Suppressing Effect of 2-Nitrobenzaldehyde on Singlet Oxygen Generation, Fatty Acid Photooxidation, and Dye-Sensitizer Degradation.

2-Nitrobenzaldehyde was found to efficiently block singlet oxygen generation in a series of different test samples upon exposure to UV and visible light under aerobic conditions. The effect of quenching singlet oxygen formation was monitored in the presence of 1, 4-diazabicyclo [2.2.2] octane (DABCO) acting as a well-known singlet oxygen scavenger. A comparison of different nitrobenzaldehyde isomers with other highly effective synthetic antioxidants used in the food industry such as butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), tert-butylhydroquinone (TBHQ) revealed that the protection of materials from singlet oxygen decreases in the order of 2-nitrobenzaldehyde > DABCO > TBHQ > 3-nitrobenzaldehyde > BHA > 4-nitrobenzaldehyde > BHT. Upon addition of 2-nitrobenzaldehyde, the oxidation of fatty acids and the degradation of photosensitizers was found to be considerably diminished, which indicates that the presence of 2-nitrobenzaldehyde has a significant protective influence by restricting the singlet oxygen generation and photodegradation of dyes. Moreover, the compound turned out to display its highly suppressing effects on typical singlet oxygen-dependent reactions, such as fatty acid photooxidation and dye photosensitizer degradation, in a rather broad spectral region covering wavelengths from 300 nm (UV-B) to 575 nm (close to the maximum of ambient solar radiation).

The Effect of Magnetite Nanoparticles and Bacteria on the Activity of NADPH-Oxidase and Myeloperoxidase in Neutrophils of Human Blood

Differences in the oxygen-dependent reactions of neutrophil granulocytes (NGs) depending on the nature of the agent affecting the cells were revealed. In vitro magnetite nanoparticles (MNPs) cause suppression of the NADPH–oxidase activity of NGs, which manifests itself in falling rates of reactions (NBT test) both with the effect of MNPs on NGs alone and a combined effect (MNPs and zymosan), as well as in the reduction of the index of activation (IA) and functional reserve of neutrophils (FRN). However, the introduction of MNPs dose-dependently stimulates the activity of myeloperoxidase (MPO). Gram-positive (S. aureus 2879 M) and gram-negative (E. coli 321) bacteria caused a respiratory burst of neutrophils, which manifested itself in a significant increase in the number of NBT-positive cells in single and combined influences (bacteria and zymosan). The lack of differences in the reaction of cells on opsonized and nonopsonized bacteria and the decrease in IA and FRN suggest that NGs are at the maximum level of functionality. Both strains of bacteria caused activation of the MPO.

Online Measurement of Oxygen-Dependent Enzyme Reaction Kinetics.

As the application of biocatalysis to complement conventional chemical and catalytic approaches continues to expand, an increasing number of reactions involve poorly water-soluble substrates. At required industrial concentrations necessary for industrial implementation, this frequently leads to heterogeneous reaction mixtures composed of multiple phases. Such systems are challenging to sample, and therefore, it is problematic to measure representative component concentrations. Herein, an online method for following the progress of oxygen-dependent reactions through accurate measurement of the oxygen mass balance in the gas phase of a reactor is demonstrated and validated. The method was successfully validated and demonstrated by using two model reactions: firstly, the oxidation of glucose by glucose oxidase and, secondly, the Baeyer-Villiger oxidation of macrocyclic ketones to lactones. Initial reaction rate constants and time-course progressions calculated from the oxygen mass balance were validated against conventional online methods of dissolved oxygen tension and pH titration measurements. A feasible operating window and the sensitivity to dynamic changes of reaction rates were established by controlling oxygen transfer through the operating parameters of the reactor. Such kinetic data forms the basis for reaction characterisation, from which bottlenecks may be made evident and directed improvement strategies can be identified and implemented.

Relative Propensities of Cytochrome c Oxidase and Cobalt Corrins for Reaction with Cyanide and Oxygen: Implications for Amelioration of Cyanide Toxicity.

In aqueous media at neutral pH, the binding of two cyanide molecules per cobinamide can be described by two formation constants, Kf1 = 1.1 (±0.6) × 105 M-1 and Kf2 = 8.5 (±0.1) × 104 M-1, or an overall cyanide binding constant of ∼1 × 1010 M-2. In comparison, the cyanide binding constants for cobalamin and a fully oxidized form of cytochrome c oxidase, each binding a single cyanide anion, were found to be 7.9 (±0.5) × 104 M-1 and 1.6 (±0.2) × 107 M-1, respectively. An examination of the cyanide-binding properties of cobinamide at neutral pH by stopped-flow spectrophotometry revealed two kinetic phases, rapid and slow, with apparent second-order rate constants of 3.2 (±0.5) × 103 M-1 s-1 and 45 (±1) M-1 s-1, respectively. Under the same conditions, cobalamin exhibited a single slow cyanide-binding kinetic phase with a second-order rate constant of 35 (±1) M-1 s-1. All three of these processes are significantly slower than the rate at which cyanide is bound by complex IV during enzyme turnover (>106 M-1 s-1). Overall, it can be understood from these findings why cobinamide is a measurably better cyanide scavenger than cobalamin, but it is unclear how either cobalt corrin can be antidotal toward cyanide intoxication as neither compound, by itself, appears able to out-compete cytochrome c oxidase for available cyanide. Furthermore, it has also been possible to unequivocally show in head-to-head comparison assays that the enzyme does indeed have greater affinity for cyanide than both cobalamin and cobinamide. A plausible resolution of the paradox that both cobalamin and cobinamide clearly are antidotal toward cyanide intoxication, involving the endogenous auxiliary agent nitric oxide, is suggested. Additionally, the catalytic consumption of oxygen by the cobalt corrins is demonstrated and, in the case of cobinamide, the involvement of cytochrome c when present. Particularly in the case of cobinamide, these oxygen-dependent reactions could potentially lead to erroneous assessment of the ability of the cyanide scavenger to restore the activity of cyanide-inhibited cytochrome c oxidase.

Oxidative stress and mitochondrial dysfunction as key players in neurological disorders of childhood

An unavoidable consequence of aerobic respiration is the generation of reactive oxygen species (ROS). ROS is a collective term that includes both oxygen radicals and certain oxidizing agents that are easily converted into radicals. They can be produced from both endogenous and exogenous substances. ROS play a dual role in biological systems, since they can be either harmful or beneficial to living systems. They can be considered a double-edged sword: oxygen-dependent reactions and aerobic respiration have significant advantages but overproduction of ROS, a consequence of oxygen-dependent reactions, has the potential to cause damage. Overproduction of ROS may negatively impact neonatal growth and contribute to the aetiology of many developmental disorders. During mitochondrial respiration, an inability to neutralize reactive oxygen species and free radicals leads to oxidative stress. The inner membrane of the mitochondria contains a large number of free radical scavengers including glutathione, vitamin C, and vitamin E, as well as anti-oxidant enzymes such as superoxide dismutase. The brain is particularly vulnerable to free radical attack for several reasons, including exposed to high oxygen concentrations, relatively low antioxidant protection, membranes with high levels of polyunsaturated fatty acids, and high iron and ascorbate content. The brain's high energy demand is primarily supplied by oxidative phosphorylation, the major producer of free radicals. When the level of free radicals overwhelms the cellular antioxidant defense system, a deleterious condition known as oxidative stress occurs. ROS has an intimate relationship with mitochondrial function and oxidative stress is believed to result from mitochondrial dysfunction. This review highlights the role of oxidative stress and mitochondrial dysfunction as key players in the neurodevelopmental pathophysiology. The mechanisms associating these two disease states can lead to neuronal death, neuroinflammation and impairment of energy metabolism. Biomarkers related to both aspects will be highlighted to demonstrate their importance in the early diagnosis of neurodevelopmental disorders like autism, cerebral palsy and others. Treatments trials for oxidative stress or mitochondrial dysfunction using nutritional supplements and antioxidants are reviewed in order to shed light on recent strategies for the early intervention for neurodevelopmental disorders.

Oxidative stress in neonatology: Role of reactive oxygen species

Oxygen is one of the most abundant elements on Earth and plays a central role in the evolution of complex life by maintaining a homeostasis within our planet biosphere. However, oxygen can also produce toxic molecules such as reactive oxygen species (ROS). ROS is a collective term that includes both free oxygen radicals and certain oxidizing agents that are easily converted into free radicals. ROS can be produced from both endogenous and exogenous substances and play a dual role in different biological systems, either harmful or beneficial to living organisms. On one hand, oxygen-dependent reactions and aerobic respiration have significant advantages but, on the other, overproduction of ROS has the potential to cause tissue damage. Living organisms have developed different ways of counteracting the harmful effects of ROS: by preventive mechanisms; repair; and physical and antioxidant defenses. The study of the interplay of the generation of ROS and their counteracting antioxidant defense systems during fetal and neonatal life is constantly changing and still not understood fully. New studies are being carried out to define the biological roles of these highly reactive species and in-vivo maintenance of optimal oxidant/antioxidant balance in neonates. It is now known that preterm birth, itself, is associated with an increased oxidant burden, which places these infants at much higher risk of tissue injury. In this article, we discuss the biochemical and physiological aspects of ROS-mediated injury in neonates.

Oxylipins in moss development and defense.

Oxylipins are oxygenated fatty acids that participate in plant development and defense against pathogen infection, insects, and wounding. Initial oxygenation of substrate fatty acids is mainly catalyzed by lipoxygenases (LOXs) and α-dioxygenases but can also take place non-enzymatically by autoxidation or singlet oxygen-dependent reactions. The resulting hydroperoxides are further metabolized by secondary enzymes to produce a large variety of compounds, including the hormone jasmonic acid (JA) and short-chain green leaf volatiles. In flowering plants, which lack arachidonic acid, oxylipins are produced mainly from oxidation of polyunsaturated C18 fatty acids, notably linolenic and linoleic acids. Algae and mosses in addition possess polyunsaturated C20 fatty acids including arachidonic and eicosapentaenoic acids, which can also be oxidized by LOXs and transformed into bioactive compounds. Mosses are phylogenetically placed between unicellular green algae and flowering plants, allowing evolutionary studies of the different oxylipin pathways. During the last years the moss Physcomitrella patens has become an attractive model plant for understanding oxylipin biosynthesis and diversity. In addition to the advantageous evolutionary position, functional studies of the different oxylipin-forming enzymes can be performed in this moss by targeted gene disruption or single point mutations by means of homologous recombination. Biochemical characterization of several oxylipin-producing enzymes and oxylipin profiling in P. patens reveal the presence of a wider range of oxylipins compared to flowering plants, including C18 as well as C20-derived oxylipins. Surprisingly, one of the most active oxylipins in plants, JA, is not synthesized in this moss. In this review, we present an overview of oxylipins produced in mosses and discuss the current knowledge related to the involvement of oxylipin-producing enzymes and their products in moss development and defense.

A Novel “Oxygen-induced” Greening Process in a Cyanobacterial Mutant Lacking the Transcriptional Activator ChlR Involved in Low-oxygen Adaptation of Tetrapyrrole Biosynthesis

ChlR activates the transcription of the chlAII-ho2-hemN operon in response to low-oxygen conditions in the cyanobacterium Synechocystis sp. PCC 6803. Three genes in the operon encode low-oxygen-type enzymes to bypass three oxygen-dependent reactions in tetrapyrrole biosynthesis. A chlR-lacking mutant, ΔchlR, shows poor photoautotrophic growth due to low chlorophyll (Chl) content under low-oxygen conditions, which is caused by no induction of the operon. Here, we characterized the processes of etiolation of ΔchlR cells in low-oxygen conditions and the subsequent regreening of the etiolated cells upon exposure to oxygen, by HPLC, Western blotting, and low-temperature fluorescence spectra. The Chl content of the etiolated ΔchlR cells incubated under low-oxygen conditions for 7 days was only 10% of that of the wild-type with accumulation of almost all intermediates of the magnesium branch of Chl biosynthesis. Both photosystem I (PSI) and photosystem II (PSII) were significantly decreased, accompanied by a preferential decrease of antenna Chl in PSI. Upon exposure to oxygen, the etiolated ΔchlR cells resumed to produce Chl after a short lag (∼2 h), and the level at 72 h was 80% of that of the wild-type. During this novel "oxygen-induced" greening process, the PSI and PSII contents were largely increased in parallel with the increase in Chl contents. After 72 h, the PSI content reached ∼50% of the wild-type level in contrast to the full recovery of PSII. ΔchlR provides a promising alternative system to investigate the biogenesis of PSI and PSII.

321: Gender peculiarities of lipid profile and metabolic oxygen-dependent reactions in patients with acute myocardial infarction accompanied with non-alcoholic steatohepatitis

It is well known that prognosis of acute myocardial infarction (AMI) in women (taking into consideration the age correction) is more unfavorable than in men. That's why studying of pathogenetic issues of AMI in women with metabolic disorders is highly important. The purpose of the research is to determine the gender peculiarities of lipid profile changes and the state of metabolic oxygen-dependent reactions in patients with AMI accompanied with NASH. 59 men and 36 women with AMI accompanied with NASH were examined. All women had menopause. The amount of troponin, creatine phosphokinase, transaminases was determined in blood of all patients. Also the data of liver ultrasound and liver biopsy were analyzed. The state of metabolic oxygen-dependent reactions was determined by spectrophotometric method; the levels of malonic dialdehyde (MDA) and diene conjugates (DC) were analyzed. Women had higher indexes of MDA (+10,2%, p=0,028) and DC (+15,2%, p=0,009) in comparison with men. Also, a decreased activity of superoxide dismutase and ceruloplasmin was revealed in women (−12,7% and − 3,3% respectively, p<0,05). Disorders of blood lipid profile were determined in patients of both groups. However, women had reliably higher level of cholesterol (+15,4%, p=0,027), triglycerides (+22,8%, p=0,003) and low-density lipoproteins (+15,8%, p=0,015). Besides, the level of high-density lipoprotein in women was lower than in men (−17,6%, p=0,022). Women with AMI accompanied with NASH were characterized by more significant derangements of lipid profile and higher activity of metabolic oxygen-dependent reactions than men. The mentioned above changes in women were probably caused by loss of estrogen protection due to menopause. Such changes suggest the need in more intensive hypolipidemic and antioxidant therapy in women with AMI accompanied with NASH.

MarR-type Transcriptional Regulator ChlR Activates Expression of Tetrapyrrole Biosynthesis Genes in Response to Low-oxygen Conditions in Cyanobacteria

Oxygen is required for three enzyme reactions in chlorophyll and bilin biosynthesis pathways: coproporphyrinogen III oxidase (HemF), heme oxygenase (HO1), and Mg-protoporphyrin IX monomethylester cyclase (ChlA(I)). The cyanobacterium Synechocystis sp. PCC 6803 has alternative enzymes, HemN, HO2, and ChlA(II), to supply chlorophyll/bilins even under low-oxygen environments. The three genes form an operon, chlA(II)-ho2-hemN, that is induced in response to low-oxygen conditions to bypass the oxygen-dependent reactions. Here we identified a transcriptional regulator for the induction of the operon in response to low-oxygen conditions. A pseudorevertant, Δho1R, was isolated from a HO1-lacking mutant Δho1 that is lethal under aerobic conditions. Δho1R grew well even under aerobic conditions. In Δho1R, HO2 that is induced only under low-oxygen conditions was anomalously expressed under aerobic conditions to complement the loss of HO1. A G-to-C transversion in sll1512 causing the amino acid change from aspartate 35 to histidine was identified as the relevant mutation by resequencing of the Δho1R genome. Sll1512 is a MarR-type transcriptional regulator. An sll1512-lacking mutant grew poorly under low-oxygen conditions with a remarked decrease in Chl content that would be caused by the suppressed induction of the chlA(II) and hemN genes in Chl biosynthesis under low-oxygen conditions. These results demonstrated that Sll1512 is an activator in response to low-oxygen environments and that the D35H variant becomes a constitutive activator. This hypothesis was supported by a gel shift assay showing that the Sll1512-D35H variant binds to the DNA fragment upstream of the operon. We propose to name sll1512 chlR.

The Antimicrobial Defense of the Pacific Oyster, Crassostrea gigas. How Diversity may Compensate for Scarcity in the Regulation of Resident/Pathogenic Microflora

Healthy oysters are inhabited by abundant microbial communities that vary with environmental conditions and coexist with immunocompetent cells in the circulatory system. In Crassostrea gigas oysters, the antimicrobial response, which is believed to control pathogens and commensals, relies on potent oxygen-dependent reactions and on antimicrobial peptides/proteins (AMPs) produced at low concentrations by epithelial cells and/or circulating hemocytes. In non-diseased oysters, hemocytes express basal levels of defensins (Cg-Defs) and proline-rich peptides (Cg-Prps). When the bacterial load dramatically increases in oyster tissues, both AMP families are driven to sites of infection by major hemocyte movements, together with bactericidal permeability/increasing proteins (Cg-BPIs) and given forms of big defensins (Cg-BigDef), whose expression in hemocytes is induced by infection. Co-localization of AMPs at sites of infection could be determinant in limiting invasion as synergies take place between peptide families, a phenomenon which is potentiated by the considerable diversity of AMP sequences. Besides, diversity occurs at the level of oyster AMP mechanisms of action, which range from membrane lysis for Cg-BPI to inhibition of metabolic pathways for Cg-Defs. The combination of such different mechanisms of action may account for the synergistic activities observed and compensate for the low peptide concentrations in C. gigas cells and tissues. To overcome the oyster antimicrobial response, oyster pathogens have developed subtle mechanisms of resistance and evasion. Thus, some Vibrio strains pathogenic for oysters are equipped with AMP-sensing systems that trigger resistance. More generally, the known oyster pathogenic vibrios have evolved strategies to evade intracellular killing through phagocytosis and the associated oxidative burst.

A Novel Testosterone Catabolic Pathway in Bacteria

Forty years ago, Coulter and Talalay (A. W. Coulter and P. Talalay, J. Biol. Chem. 243:3238-3247, 1968) established the oxygenase-dependent pathway for the degradation of testosterone by aerobes. The oxic testosterone catabolic pathway involves several oxygen-dependent reactions and is not available for anaerobes. Since then, a variety of anaerobic bacteria have been described for the ability to degrade testosterone in the absence of oxygen. Here, a novel, oxygenase-independent testosterone catabolic pathway in such organisms is described. Steroidobacter denitrificans DSMZ18526 was shown to be capable of degrading testosterone in the absence of oxygen and was selected as the model organism in this study. In a previous investigation, we identified the initial intermediates involved in an anoxic testosterone catabolic pathway, most of which are identical to those of the oxic pathway demonstrated in Comamonas testosteroni. In this study, five additional intermediates of the anoxic pathway were identified. We demonstrated that subsequent steps of the anoxic pathway greatly differ from those of the established oxic pathway, which suggests that a novel pathway for testosterone catabolism is present. In the proposed anoxic pathway, a reduction reaction occurs at C-4 and C-5 of androsta-1,4-diene-3,17-dione, the last common intermediate of both the oxic and anoxic pathways. After that, a novel hydration reaction occurs and a hydroxyl group is thus introduced to the C-1α position of C(19)steroid substrates. To our knowledge, an enzymatic hydration reaction occurring at the A ring of steroid compounds has not been reported before.

Oxygen toxicity: chemistry and biology of reactive oxygen species

Oxygen has a central role in the evolution of complex life on Earth mainly because of the biochemical symmetry of oxygenic photosynthesis and aerobic respiration that can maintain homeostasis within our planet biosphere. Oxygen can also produce toxic molecules, reactive oxygen species (ROS). ROS is a collective term that includes both oxygen radicals and certain oxidizing agents that are easily converted into radicals. They can be produced from both endogenous and exogenous substances. ROS play a dual role in biological systems, since they can be either harmful or beneficial to living systems. They can be considered a double-edged sword because on the one hand oxygen-dependent reactions and aerobic respiration have significant advantages but, on the other, overproduction of ROS has the potential to cause damage.

Identification of an Atypical Membrane Protein Involved in the Formation of Protein Disulfide Bonds in Oxygenic Photosynthetic Organisms

The evolution of oxygenic photosynthesis in cyanobacteria nearly three billion years ago provided abundant reducing power and facilitated the elaboration of numerous oxygen-dependent reactions in our biosphere. Cyanobacteria contain an internal thylakoid membrane system, the site of photosynthesis, and a typical Gram-negative envelope membrane system. Like other organisms, the extracytoplasmic space in cyanobacteria houses numerous cysteine-containing proteins. However, the existence of a biochemical system for disulfide bond formation in cyanobacteria remains to be determined. Extracytoplasmic disulfide bond formation in non-photosynthetic organisms is catalyzed by coordinated interaction between two proteins, a disulfide carrier and a disulfide generator. Here we describe a novel gene, SyndsbAB, required for disulfide bond formation in the extracytoplasmic space of cyanobacteria. The SynDsbAB orthologs are present in most cyanobacteria and chloroplasts of higher plants with fully sequenced genomes. The SynDsbAB protein contains two distinct catalytic domains that display significant similarity to proteins involved in disulfide bond formation in Escherichia coli and eukaryotes. Importantly, SyndsbAB complements E. coli strains defective in disulfide bond formation. In addition, the activity of E. coli alkaline phosphatase localized to the periplasm of Synechocystis 6803 is dependent on the function of SynDsbAB. Deletion of SyndsbAB in Synechocystis 6803 causes significant growth impairment under photoautotrophic conditions and results in hyper-sensitivity to dithiothreitol, a reductant, whereas diamide, an oxidant had no effect on the growth of the mutant strains. We conclude that SynDsbAB is a critical protein for disulfide bond formation in oxygenic photosynthetic organisms and required for their optimal photoautotrophic growth.

Die Physiologie der Wundheilung

Wound healing is a complex biological phenomenon. A variety of cellular and biochemical events take place designed to achieve tissue integrity following injury. Even though hypoxia caused by the damaged microvasculature is an important initial stimulus of the healing cascade, adequate tissue perfusion and oxygenation is an absolute pre-requisite for a successful repair since essential wound healing mechanisms such as collagen deposition and bactericidal defence are oxygen dependent reactions. Based upon these findings, there are several ways to overcome the obstacle of tissue hypoxia in clinical practice. Supplemental oxygen is capable of increasing tissue oxygen tension. Following surgery, pain, cold and to little fluids are the main issues of vasoconstriction, impaired tissue perfusion and oxygenation as well. However, all these parameters must be corrected at the same time because any one is sufficient to cause maximal vasoconstriction. A well hydrated, pain free and warm patient should be the main goal of peri- and postoperative surgical care.

Unusual reactions involved in anaerobic metabolism of phenolic compounds

Aerobic bacteria use molecular oxygen as a common co-substrate for key enzymes of aromatic metabolism. In contrast, in anaerobes all oxygen-dependent reactions are replaced by a set of alternative enzymatic processes. The anaerobic degradation of phenol to a non-aromatic product involves enzymatic processes that are uniquely found in the aromatic metabolism of anaerobic bacteria: (i) ATP-dependent phenol carboxylation to 4-hydroxybenzoate via a phenylphosphate intermediate (biological Kolbe-Schmitt carboxylation); (ii) reductive dehydroxylation of 4-hydroxybenzoyl-CoA to benzoyl-CoA; and (iii) ATP-dependent reductive dearomatization of the key intermediate benzoyl-CoA in a 'Birch-like' reduction mechanism. This review summarizes the results of recent mechanistic studies of the enzymes involved in these three key reactions.