Ein neues Konzept für das bedarfsgerechte Informations- und Wissensmanagement in Unternehmenskooperationen der Multimaterial-Mikrosystemtechnik

In der Arbeit wird ein Konzept zur Kooperation mittelständischer Unternehmen der Mikrosystemtechnik vorgestellt, das neben der zeitlichen Entwicklung der Kooperationsintensität auch ein zeitabhängiges, die Kooperations-Kernprozesse unterstützendes Informations- und Wissensmanagement adressiert. Die Arbeit identifiziert drei Entwicklungsphasen, insbesondere unter dem Aspekt der verteilten Produktentwicklung. Für die Kooperationsprozesse werden bedarfsgerechte Softwarewerkzeuge vorgestellt

MerA functions as a hypothiocyanous acid reductase and defense mechanism in Staphylococcus aureus

The major pathogen Staphylococcus aureus has to cope with host-derived oxidative stress to cause infections in humans. Here, we report that S. aureus tolerates high concentrations of hypothiocyanous acid (HOSCN), a key antimicrobial oxidant produced in the respiratory tract. We discovered that the flavoprotein disulfide reductase (FDR) MerA protects S. aureus from this oxidant by functioning as a HOSCN reductase, with its deletion sensitizing bacteria to HOSCN. Crystal structures of homodimeric MerA (2.4 Å) with a Cys43–Cys48 intramolecular disulfide, and reduced MerACys43S (1.6 Å) showed the FAD cofactor close to the active site, supporting that MerA functions as a group I FDR. MerA is controlled by the redox-sensitive repressor HypR, which we show to be oxidized to intermolecular disulfides under HOSCN stress, resulting in its inactivation and derepression of merA transcription to promote HOSCN tolerance. Our study highlights the HOSCN tolerance of S. aureus and characterizes the structure and function of MerA as a major HOSCN defense mechanism. Crippling the capacity to respond to HOSCN may be a novel strategy for treating S. aureus infections

Measuring chlorine bleach in biology and medicine

Background: Chlorine bleach, or hypochlorous acid, is the most reactive two-electron oxidant produced in appreciable amounts in our bodies. Neutrophils are the main source of hypochlorous acid. These champions of the innate immune system use it to fight infection but also direct it against host tissue in inflammatory diseases. Neutrophils contain a rich supply of the enzyme myeloperoxidase. It uses hydrogen peroxide to convert chloride to hypochlorous acid. Scope of review: We give a critical appraisal of the best methods to measure production of hypochlorous acid by purified peroxidases and isolated neutrophils. Robust ways of detecting it inside neutrophil phagosomes where bacteria are killed are also discussed. Special attention is focused on reaction-based fluorescent probes but their visual charm is tempered by stressing their current limitations. Finally, the strengths and weaknesses of biomarker assays that capture the footprints of chlorine in various pathologies are evaluated. Major conclusions: Detection of hypochlorous acid by purified peroxidases and isolated neutrophils is best achieved by measuring accumulation of taurine chloramine. Formation of hypochlorous acid inside neutrophil phagosomes can be tracked using mass spectrometric analysis of 3-chlorotyrosine and methionine sulfoxide in bacterial proteins, or detection of chlorinated fluorescein on ingestible particles. Reaction-based fluorescent probes can also be used to monitor hypochlorous acid during phagocytosis. Specific biomarkers of its formation during inflammation include 3-chlorotyrosine, chlorinated products of plasmalogens, and glutathione sulfonamide. General significance: These methods should bring new insights into how chlorine bleach is produced by peroxidases, reacts within phagosomes to kill bacteria, and contributes to inflammation. This article is part of a Special Issue entitled Current methods to study reactive oxygen species - pros and cons and biophysics of membrane proteins. Guest Editor: Christine Winterbourn

Oxidation of bacillithiol during killing of Staphylococcus aureus USA300 inside neutrophil phagosomes

Targeting immune evasion tactics of pathogenic bacteria may hold the key to treating recalcitrant bacterial infections. Staphylococcus aureus produces bacillithiol (BSH), its major low-molecular-weight thiol, which is thought to protect this opportunistic human pathogen against the bombardment of oxidants inside neutrophil phagosomes. Here, we show that BSH was oxidized when human neutrophils phagocytosed S. aureus, but provided limited protection to the bacteria. We used mass spectrometry to measure the oxidation of BSH upon exposure of S. aureus USA300 to either a bolus of hypochlorous acid (HOCl) or a flux generated by the neutrophil enzyme myeloperoxidase. Oxidation of BSH and loss of bacterial viability were strongly correlated (r = 0.99, p < 0.001). BSH was fully oxidized after exposure of S. aureus to lethal doses of HOCl. However, there was no relationship between the initial BSH levels and the dose of HOCl required for bacterial killing. In contrast to the HOCl systems, only 50% of total BSH was oxidized when neutrophils killed the majority of phagocytosed bacteria. Oxidation of BSH was decreased upon inhibition of myeloperoxidase, implicating HOCl in phagosomal BSH oxidation. A BSH-deficient S. aureus USA300 mutant was slightly more susceptible to treatment with either HOCl or ammonia chloramine, or to killing within neutrophil phagosomes. Collectively, our data show that myeloperoxidase-derived oxidants react with S. aureus inside neutrophil phagosomes, leading to partial BSH oxidation, and contribute to bacterial killing. However, BSH offers only limited protection against the neutrophil's multifaceted killing mechanisms

Uric acid and thiocyanate as competing substrates of lactoperoxidase

The physiological function of urate is poorly understood. It may act as a danger signal, an antioxidant, or a substrate for heme peroxidases. Whether it reacts sufficiently rapidly with lactoperoxidase (LPO) to act as a physiological substrate remains unknown. LPO is a mammalian peroxidase that plays a key role in the innate immune defense by oxidizing thiocyanate to the bactericidal and fungicidal agent hypothiocyanite. We now demonstrate that urate is a good substrate for bovine LPO

A myeloperoxidase precursor, promyeloperoxidase, is present in human plasma and elevated in cardiovascular disease patients

Myeloperoxidase (MPO)-derived oxidants have emerged as a key contributor to tissue damage in inflammatory conditions such as cardiovascular disease. Pro-myeloperoxidase (pro-MPO), an enzymatically active precursor of myeloperoxidase (MPO), is known to be secreted from cultured bone marrow and promyelocytic leukemia cells, but evidence for the presence of pro-MPO in circulation is lacking. In the present study, we used a LC-MS/MS in addition to immunoblot analyses to show that pro-MPO is present in human blood plasma. Furthermore, we found that pro-MPO was more frequently detected in plasma from patients with myocardial infarction compared to plasma from control donors. Our study suggests that in addition to mature MPO, circulating pro-MPO may cause oxidative modifications of proteins thereby contributing to cardiovascular disease

Post-translational regulation of macrophage migration inhibitory factor: Basis for functional fine-tuning

Macrophage migration inhibitory factor (MIF) is a chemokine-like protein and an important mediator in the inflammatory response. Unlike most other pro-inflammatory cytokines, a number of cell types constitutively express MIF and secretion occurs from preformed stores. MIF is an evolutionarily conserved protein that shows a remarkable functional diversity, including specific binding to surface CD74 and chemokine receptors and the presence of two intrinsic tautomerase and oxidoreductase activities. Several studies have shown that MIF is subject to post-translational modification, particularly redox-dependent modification of the catalytic proline and cysteine residues. In this review, we summarize and discuss MIF post-translational modifications and their effects on the biological properties of this protein. We propose that the redox-sensitive residues in MIF will be modified at sites of inflammation and that this will add further depth to the functional diversity of this intriguing cytokine