Redox signalling and detection of protein oxidation by mass spectrometry

It is now recognised that redox control of proteins plays an important role in many signalling pathways both in health and disease. Proteins can undergo a wide variety of oxidative post-translational modifications (oxPTM); while the reversible modifications are thought to be most important in physiological processes, non-reversible oxPTM may contribute to pathological situations and disease. The oxidant is also important in determining the type of oxPTM (chlorination, nitration, etc.), and the susceptibilities of residues vary depending on their structural location. The best characterized oxPTMs involved in signalling modulation are partial oxidations of cysteine to the disulfide, glutathionylated or sulfenic acid forms, but there is increasing evidence that specific oxidations of methionine and tyrosine may have some biological roles. Well understood examples of oxidative regulation include protein tyrosine phosphatases, e.g. PTP1B/C, and members of the MAPK pathways such as MEKK1 and ASK1. Transcription factors such as NFkB and Nrf-2 are also regulated by redox-active cysteines. Improved methods for analysing specific oxPTMs in biological samples are critical for understanding the physiological and pathological roles of these changes, and tandem or MS3 mass spectrometry techniques interfaced with nano-LC separation are being now used. MS3 fragmentation markers for a variety of oxidized residues including tyrosine, tryptophan and proline have been identified, and a precursor ion scanning method that allows the selective identification of these oxPTMs in complex samples has been developed. Such advances in technology offer potential for biomarker development, disease diagnosis and understanding pathology.

Chemistry of phospholipid oxidation

The oxidation of lipids has long been a topic of interest in biological and food sciences, and the fundamental principles of non-enzymatic free radical attack on phospholipids are well established, although questions about detail of the mechanisms remain. The number of end products that are formed following the initiation of phospholipid peroxidation is large, and is continually growing as new structures of oxidized phospholipids are elucidated. Common products are phospholipids with esterified isoprostane-like structures and chain-shortened products containing hydroxy, carbonyl or carboxylic acid groups; the carbonyl-containing compounds are reactive and readily form adducts with proteins and other biomolecules. Phospholipids can also be attacked by reactive nitrogen and chlorine species, further expanding the range of products to nitrated and chlorinated phospholipids. Key to understanding the mechanisms of oxidation is the development of advanced and sensitive technologies that enable structural elucidation. Tandem mass spectrometry has proved invaluable in this respect and is generally the method of choice for structural work. A number of studies have investigated whether individual oxidized phospholipid products occur in vivo, and mass spectrometry techniques have been instrumental in detecting a variety of oxidation products in biological samples such as atherosclerotic plaque material, brain tissue, intestinal tissue and plasma, although relatively few have achieved an absolute quantitative analysis. The levels of oxidized phospholipids in vivo is a critical question, as there is now substantial evidence that many of these compounds are bioactive and could contribute to pathology. The challenges for the future will be to adopt lipidomic approaches to map the profile of oxidized phospholipid formation in different biological conditions, and relate this to their effects in vivo. This article is part of a Special Issue entitled: Oxidized phospholipids-their properties and interactions with proteins.

High temperature gas cleaning for advanced syngas utilization:Paper # 95

Biofuels and chemicals from biomass mean the gasification of biogenic feedstocks and the synthesis via methanol, dimethylester (DME) or Fischer-Tropsch products. To prevent the sensitive synthesis catalysts from poisoning the syngas must be free of tar and particulates. The trace concentrations of S-, C1-, N-species, alkali and heavy metals must be of the order of a few ppb. Moreover maximum conversion efficiency will be achieved performing the gas cleaning above the synthesis conditions. The concept of an innovative dry HTHP syngas cleaning is presented. Based on the HT particle filtration and suitable sorption and catalysis processes for the relevant contaminants a total concept will be derived, which leads to a syngas quality required for synthesis catalysts in only 2 combined stages. The experimental setup for the HT gas cleaning behind the 60 kWtherm entrained flow gasifier REGA of the institute is described. Results from HT filter experiments in pilot scale are presented. The performance of 2 natural minerals for HC1 and H2S sorption is discussed with respect to the parameters temperature, surface and residence time. Results from lab scale investigations on low temperature tar catalysts' performance (commercial and proprietary development) are discussed finally.

Reaction-driven surface restructuring and selectivity control in allylic alcohol catalytic aerobic oxidation over Pd

Synchronous, time-resolved DRIFTS/MS/XAS cycling studies of the vapor-phase selective aerobic oxidation of crotyl alcohol over nanoparticulate Pd have revealed surface oxide as the desired catalytically active phase, with dynamic, reaction-induced Pd redox processes controlling selective versus combustion pathways.

Shopfloor attitudes towards advanced manufacturing technology:the changing focus of industrial conflict?

The implementation of advanced manufacturing technology (AMT) in manufacturing organisations is increasing. In many cases the introduction of AMT has been associated with conflict between management and workers. This appears to be due to the potential for AMT to have a de-skilling effect upon job content and, in some instances, leading to job losses. In reality, fears concerning both these issues have reduced and consequently there has been a change away from conflict between management and workers to divisions amongst shopfloor operators. The paper explores some of the processes involved in this change within the context of an engineering case study. More specifically, it is shown that when AMT was introduced into a machining workshop, traditional conflict between management and operators was soon replaced by negative feelings between users and non-users of AMT. The implications of industrial relations suggest the need for more care and attention to the human side of work organisation when implementing new manufacturing technology.

Servitization of the manufacturing firm:exploring the operations practices and technologies that deliver advanced services

Purpose: This paper aims to explore practices and technologies successfully servitised manufacturers employ in the delivery of advanced services. Design/methodology/approach: A case study methodology is applied across four manufacturing organisations successful in servitization. Through interviews with personnel across host manufacturers, their partners, and key customers, extensive data are collected about service delivery systems. Analyses identify convergence in their practices and technologies. Findings: Six distinct technologies and practices are revealed: facilities and their location, micro-vertical integration and supplier relationships, information and communication technologies (ICTs), performance measurement and value demonstration, people deployment and their skills, and business processes and customer relationships. These are then combined in an integrative framework that illustrates how operations are configured to successfully deliver advanced services. Research limitations/implications: The analyses are reductive and rationalising. Future studies could identify other technologies and practices. Case study as a method is inherently limited in the extent to which findings can be generalised. Practical implications: Awareness and interest in servitization is growing, yet adoption of a servitization strategy requires particular organisational capabilities on the part of the manufacturer. This study identifies technologies and practices that underpin these capabilities. Originality/value: This paper contributes to the understanding of the servitization process and, in particular, the implications to broader operations of the firm. © Emerald Group Publishing Limited.

A review of advanced catalyst development for Fischer-Tropsch synthesis of hydrocarbons from biomass derived syn-gas

Fischer-Tropsch synthesis (FTS) is a process which converts syn-gas (H2 and CO) to synthetic liquid fuels and valuable chemicals. Thermal gasification of biomass represents a convenient route to produce syn-gas from intractable materials particularly those derived from waste that are not cost effective to process for use in biocatalytic or other milder catalytic processes. The development of novel catalysts with high activity and selectivity is desirable as it leads to improved quality and value of FTS products. This review paper summarises recent developments in FT-catalyst design with regards to optimising catalyst activity and selectivity towards synthetic fuels. © 2014 the Partner Organisations.

Proteomic analysis of phosphorylation, oxidation and nitrosylation in signal transduction

Signal transduction pathways control cell fate, survival and function. They are organized as intricate biochemical networks which enable biochemical protein activities, crosstalk and subcellular localization to be integrated and tuned to produce highly specific biological responses in a robust and reproducible manner. Post translational Modifications (PTMs) play major roles in regulating these processes through a wide variety of mechanisms that include changes in protein activities, interactions, and subcellular localizations. Determining and analyzing PTMs poses enormous challenges. Recent progress in mass spectrometry (MS) based proteomics have enhanced our capability to map and identify many PTMs. Here we review the current state of proteomic PTM analysis relevant for signal transduction research, focusing on two areas: phosphorylation, which is well established as a widespread key regulator of signal transduction; and oxidative modifications, which from being primarily viewed as protein damage now start to emerge as important regulatory mechanisms.

Servitization capabilities for advanced services:a multi-actor perspective

Servitization is the process by which manufacturers add services to their product offerings and even replace products with services. The capabilities necessary to develop and deliver advanced services as part of servitization are often discussed in the literature from the manufacturer’s perspective, e.g., having a service-focused culture or the ability to sell solutions. Recent research has acknowledged the important role of customers and, to a lesser extent, other actors (e.g., intermediaries) in bringing about successful servitization, particularly for use-oriented and results-oriented advanced services. The objective of this study is to identify the capabilities required to successful develop advanced services as part of servitization by considering the perspective of manufacturers, intermediaries and customers. This study involved interviews with 33 managers in 28 large UK-based companies from these three groups, about servitization capabilities. The findings suggest that there are eight broad capabilities that are important for advanced services; 1) personnel with expertise and deep technical product knowledge, 2) methodologies for improving operational processes, helping to manage risk and reduce costs, 3) the evolution from being a product- focused manufacturer to embracing a services culture, 4) developing trusting relationships with other actors in the network to support the delivery of advanced services, 5) new innovation activities focused on financing contracts (e.g., ‘gain share’) and technology implementation (e.g., Web-based applications), 6) customer intimacy through understanding their business challenges in order to develop suitable solutions, 7) extensive infrastructure (e.g., personnel, service centres) to deliver a local service, and 8) the ability to tailor service offerings to each customer’s requirements and deliver these responsively to changing needs. The capabilities required to develop and deliver advanced services align to a need to enhance the operational performance of supplied products throughout their lifecycles and as such require greater investment than the capabilities for base and intermediate services.

Aggregation and caking processes of granular materials:continuum model and simulation with application to sugar

Aggregation and caking of particles are common severe problems in many operations and processing of granular materials, where granulated sugar is an important example. Prevention of aggregation and caking of granular materials requires a good understanding of moisture migration and caking mechanisms. In this paper, the modeling of solid bridge formation between particles is introduced, based on moisture migration of atmospheric moisture into containers packed with granular materials through vapor evaporation and condensation. A model for the caking process is then developed, based on the growth of liquid bridges (during condensation), and their hardening and subsequent creation of solid bridges (during evaporation). The predicted caking strengths agree well with some available experimental data on granulated sugar under storage conditions.

Reversible oxidation of phosphatase and tensin homolog (PTEN) alters its interactions with signaling and regulatory proteins

Phosphatase and tensin homolog (PTEN) is involved in a number of different cellular processes including metabolism, apoptosis, cell proliferation and survival. It is a redox-sensitive dual-specificity protein phosphatase that acts as a tumor suppressor by negatively regulating the PI3K/Akt pathway. While direct evidence of redox regulation of PTEN downstream signaling has been reported, the effect of PTEN redox status on its protein-protein interactions is poorly understood. PTEN-GST in its reduced and a DTT-reversible H2O2-oxidized form was immobilized on a glutathione-sepharose support and incubated with cell lysate to capture interacting proteins. Captured proteins were analyzed by LC-MSMS and comparatively quantified using label-free methods. 97 Potential protein interactors were identified, including a significant number that are novel. The abundance of fourteen interactors was found to vary significantly with the redox status of PTEN. Altered binding to PTEN was confirmed by affinity pull-down and Western blotting for Prdx1, Trx, and Anxa2, while DDB1 was validated as a novel interactor with unaltered binding. These results suggest that the redox status of PTEN causes a functional variation in the PTEN interactome. The resin capture method developed had distinct advantages in that the redox status of PTEN could be directly controlled and measured.

Capabilities for advanced services:a multi-actor perspective

Servitization involves manufacturers developing service offerings to grow revenue and profit. Advanced services, in particular, can facilitate a more service-focused organization and impact customers' business processes significantly. However, approaches to servitization are often discussed solely from the manufacturer's perspective; overlooking the role of other network actors. Adopting a multi-actor perspective, this study investigates manufacturer, intermediary and customer perspectives to identify complementary and competing capabilities within a manufacturer's downstream network, required for advanced services. Interviews were conducted with 24 senior executives in 19 UK-based manufacturers, intermediaries and customers across multiple sectors. The study identified six key business activities, within which advanced services capabilities were grouped. The unique and critical capabilities for advanced services for each actor were identified as follows: manufacturers; the need to balance product and service innovation, developing customer-focused through-life service methodologies and having distinct, yet synergistic product and service cultures; intermediaries, the coordination and integration of third party products/services; customers, co-creating innovation and having processes supporting service outsourcing. The study is unique in highlighting the distinct roles of different actors in the provision of advanced services and shows that they can only be developed and delivered by the combination of complex interconnected capabilities found within a network.

Advanced predictive-analysis-based decision support for collaborative logistics networks

Purpose – The purpose of this paper is to examine challenges and potential of big data in heterogeneous business networks and relate these to an implemented logistics solution. Design/methodology/approach – The paper establishes an overview of challenges and opportunities of current significance in the area of big data, specifically in the context of transparency and processes in heterogeneous enterprise networks. Within this context, the paper presents how existing components and purpose-driven research were combined for a solution implemented in a nationwide network for less-than-truckload consignments. Findings – Aside from providing an extended overview of today’s big data situation, the findings have shown that technical means and methods available today can comprise a feasible process transparency solution in a large heterogeneous network where legacy practices, reporting lags and incomplete data exist, yet processes are sensitive to inadequate policy changes. Practical implications – The means introduced in the paper were found to be of utility value in improving process efficiency, transparency and planning in logistics networks. The particular system design choices in the presented solution allow an incremental introduction or evolution of resource handling practices, incorporating existing fragmentary, unstructured or tacit knowledge of experienced personnel into the theoretically founded overall concept. Originality/value – The paper extends previous high-level view on the potential of big data, and presents new applied research and development results in a logistics application.

The redoxomics of PTEN: walking a fine line between damage and signaling:mass spectrometry-based approaches to study the effect of oxidation on PTEN function, structure, and protein-protein interactions

The research described in this PhD thesis focuses on proteomics approaches to study the effect of oxidation on the modification status and protein-protein interactions of PTEN, a redox-sensitive phosphatase involved in a number of cellular processes including metabolism, apoptosis, cell proliferation, and survival. While direct evidence of a redox regulation of PTEN and its downstream signaling has been reported, the effect of cellular oxidative stress or direct PTEN oxidation on PTEN structure and interactome is still poorly defined. In a first study, GST-tagged PTEN was directly oxidized over a range of hypochlorous acid (HOCl) concentration, assayed for phosphatase activity, and oxidative post-translational modifications (oxPTMs) were quantified using LC-MS/MS-based label-free methods. In a second study, GSTtagged PTEN was prepared in a reduced and reversibly H2O2-oxidized form, immobilized on a resin support and incubated with HCT116 cell lysate to capture PTEN interacting proteins, which were analyzed by LC-MS/MS and comparatively quantified using label-free methods. In parallel experiments, HCT116 cells transfected with a GFP-tagged PTEN were treated with H2O2 and PTENinteracting proteins immunoprecipitated using standard methods. Several high abundance HOCl-induced oxPTMs were mapped, including those taking place at amino acids known to be important for PTEN phosphatase activity and protein-protein interactions, such as Met35, Tyr155, Tyr240 and Tyr315. A PTEN redox interactome was also characterized, which identified a number of PTEN-interacting proteins that vary with the reversible inactivation of PTEN caused by H2O2 oxidation. These included new PTEN interactors as well as the redox proteins peroxiredoxin-1 (Prdx1) and thioredoxin (Trx), which are known to be involved in the recycling of PTEN active site following H2O2-induced reversible inactivation. The results suggest that the oxidative modification of PTEN causes functional alterations in PTEN structure and interactome, with fundamental implications for the PTEN signaling role in many cellular processes, such as those involved in the pathophysiology of disease and ageing.

eBEST platform a KKV-klaszterek operatív és együttműködési folyamatainak támogatására (eBEST platform for supporting operational and cooperative processes of SME's clusters)

Az Európai Bizottság jelentése szerint a magyar kis- és középvállalkozások helyzete 2005 óta stagnál. Bár ezek a vállalkozások adják a magyar vállalkozások 99%-át, mégis a közbeszerzési, valamint a növekvő piacokhoz való hozzáférés terén számos akadállyal kerülnek szembe. Az eBEST projekten (Empowering Business Ecosystems of Small Service Enterprises to Face the Economic Crisis) belül kialakított platform olyan funkcionalitással bír, ami mindamellett, hogy lehetővé teszi a vállalkozások szervezett csoportokba, azaz ökoszisztémákba rendeződését, hozzá tud járulni a fogyasztói igények kielégítése érdekében létrejövő ellátási lánc, illetve egyedi folyamatok mentén fellépő információszerzési, kommunikációs vagy együttműködési akadályok lebontásához. ____ It is widely recognised that the most important factor for increasing the productivity of small companies is a deep adoption of computer-based applications and services. The FP7 SME eBEST project proposed a new operational environment specifically conceived for net worked small companies, supported by an advanced suite of ICT services, the eBEST platform. The paper aims at presenting the projects achievements that are validated by a number of company clusters of different EU countries and industry sectors. The general objectives of the eBEST project are attracting customers to work with the clustered companies, facilitating companies to collaborate with each other, and enabling associations to foster the devised innovation.