Grâce aux récents progrès en terme d'instrumentation analytique, la protéomique, en tant que science qui étudie le protéome d'un organisme ou d'un milieu, a connu un véritable tournant et a permis d'étendre le champ des connaissances sur le fonctionnement du vivant dans son ensemble (structure, fonction, métabolisme, dynamisme). Néanmoins, l'étude des protéomes représente un challenge pour de nombreux biologistes, chimistes et biochimistes, en raison notamment de la complexité des échantillons étudiés. De nombreux protocoles analytiques ont d'ores et déjà été développés. Cependant, dans leur ensemble, ces stratégies sont relativement longues et multi-étapes et le plus souvent ciblées sur une protéine donnée.Dans ce contexte, ce travail de thèse a pour objectif de simplifier les protocoles expérimentaux existants afin de limiter les pertes en protéines et ainsi amplifier leurs signaux au cours d'une analyse « Bottom-up » par LC-HRMS (analyse « hors gel »). Chaque étape du processus a été décortiquée et optimisée. Dans un premier temps, les paramètres UPLC-HRMS/MS ont été optimisés afin d'améliorer la détection et la quantification des protéines présentes à des concentrations très variables dans les milieux étudiés. Ensuite, une approche de purification simplifiée qui repose sur une seule et unique étape de lavage et solubilisation des protéines a été mise au point. La démonstration de son efficacité « chimique » et « biologique » a ensuite été réalisée via une étude mécanistique au cours de laquelle les changements de conformation des protéines ont été étudiés à chacune des étapes de purification proposées. Enfin, certains paramètres influençant l'extraction des protéines à partir de ces mêmes matrices ont été étudiés afin de proposer à terme un protocole d'extraction à la carte compatible avec une analyse directe par LC-HRMS/MS. / Recent advances in proteomics have been spurred by the rapid development of hybrid and/or high-resolution mass analysers (HRMS/MS). These powerful instrumentations have led to significant improvements in « Bottom-up » approach and have enabled to deepen our knowledge on the functionality of biological systems (structure, function, metabolism, dynamic, etc).Despite their high sensibilities, the potential of such instruments could be significantly lessened by an imperfect sample pre-treatment. In this context, current sample pretreatments follow multi-steps experimental workflows, which alternatively lead to low recoveries of proteins. In this line, this study aims at developing a simple and versatile strategy in order to reduce protein losses and enhance their detection in gel-free LC-MS analysis. First, an analytical method based on liquid chromatography and tandem mass spectrometry was developed to detect and quantify complex peptides mixture. Secondly, a universal, simple and fast purification approach was designed with the aim to purify protein extracts in only one-step. For this purpose, the molecular reactivity, dynamics and conformational changes of proteins at each development step were comprehensively investigated with a set of spectroscopic techniques, in order to select the best strategy. Finally, different factors influencing extraction of proteins were investigated with the goal in the long term to propose an on-demand extraction protocol for direct analysis of proteins by LC-HRMS/MS.

Digitalization has been reshaping the media landscape in recent years, often conveying an implicit promise of becoming less dependent on physical resources. At the same time, the current understanding of digital reading goes beyond dedicated e-readers or definable digital media products such as magazines or newspapers. In fact, it must be perceived as a function or service obtained from existing and ever-expanding “digital ecosystems”. There is furthermore a clear and unambiguous trend that relatively small and mobile devices are on the rise for consuming all kinds of media. Next to potentially enabling environmental gains compared to traditional paper-based media consumption, there are agreeing indications of a shift from overall electricity consumption dominated by end-user devices towards an increasing importance of less tangible data transmission networks and data centers. Therefore, a bottom-up analysis is deemed to compliment more general top-down observations and assessments. To this end, an elaborated reference scenario is proposed as to bridge the mere analytical method of Life Cycle Assessment (LCA) with behavioral aspects based on German market observations and surveys. The prevailing aim of this study is to detect environmental hot-spots and absolute impacts linked to the service of accessing text-based content via connected electronic devices. In doing so, this study takes the position that both types of media consumption – digital and paper-based - are incommensurable due to the very evident differences in provided functions, markets, and industries. Therefore, an attributional and stand-alone LCA is considered appropriate. The perceived current situation (reference scenario) evolves around substantiated estimates and assumptions concerning production of devices, use of devices as well as operation of essential data transmission network components. Looking at potential hot-spots, electricity consumption linked to data transmission could be a decisive factor for the environmental performance of digital reading. However, the actual importance of data transmission infrastructures depends on both methodological choices and a range of parameters or trends. For instance, the relative importance is shifted when more recent estimates of electricity intensities are incorporated. Depending on actual and localized electricity intensity of data transmission, the amount of data required to provide an expected function may inhibit environmental potentials of digital media consumption. Postulating average annual consumption of digital contents and assuming actual substitution of equivalent printed media products, about 50 kg CO2-equivalents. could potentially be avoided. This theoretical potential is based on the calculated global warming potential (GWP) associated with digital reading according to the reference scenario which amounts to about 29 kg CO2-equivalents. Therefore, this study supports findings from previous studies that indicated environmental benefits of digital reading. Compared to other functions or services (e.g. video/music streaming, podcasts, audio books) embedded in the same “digital ecosystems”, reading requires little amount of data. If allocation of upstream effects is based on time, the relative importance of data transmission networks could be gauged and compared by adopting a “data-to-service time” ratio. Taking the reference scenario as a starting point, a perceivable ratio for digital reading is 0.015 GB/h, including systemic inefficiencies. In contrast, streaming of high-definition video contents can easily consume 3 GB/h, a 200-fold increase. The audience of this study comprises providers of digital reading services and/or other media services as well as end-users as integral element in “digital ecosystems”. Besides, the report proposes a conceptual assessment framework which can be applied to other contemporary digital services or functions. / De senaste åren har digitalisering omformat medielandskapet, med ett implicit löfte om att minska beroendet av fysiska resurser. Dessutom finns det tydliga trender som pekar mot en ökad användning av små, mobila enheter för att konsumera alla sorters media. En uppdaterad bottom-up analys bedöms komplettera mer generella observationer och bedömningar. Om man antar årliga genomsnittliga konsumtionsmönster i Tyskland, så är tillverkningen av elektroniska slutanvändarenheter – oavsett om de är till för enskilda ändamål (e-läsare) eller om de är multifunktionella (smartphone, surfplatta) – onekligen en miljömässigt kritisk punkt för digitalt läsande. Elförbrukningen, som sker i samband med dataöverföringen, kan också vara en avgörande faktor för den övergripande miljöpåverkan av digital läsning. Dock beror den faktiska påverkan av dataöverförningsinfrastrukturer dels på metodologiska val men även på ett antal andra parametrar och trender. Genom att undersöka indikatorn för global uppvärmning kan denna studie konstatera att resultaten stödjer tidigare forskning, som redan pekar på de miljömässiga vinsterna av digitalt läsande. Målgruppen för denna studie innefattar både distributörer av digitala läs-tjänster och/eller andra media tjänster såväl som slutanvändare som ett integrerat element i ”digitala ekosystem”.