Genome analysis and genome-wide proteomics of Thermococcus gammatolerans, the most radioresistant organism known amongst the Archaea

Background  

Thermococcus gammatolerans was isolated from samples collected from hydrothermal chimneys. It is one of the most radioresistant organisms known amongst the Archaea. We report the determination and annotation of its complete genome sequence, its comparison with other Thermococcales genomes, and a proteomic analysis.     

Elucidating the biochemical overwintering adaptations of Larval Cucujus clavipes puniceus, a nonmodel organism, via high throughput proteomics

Cucujus clavipes puniceus (C.c.p.) is a nonmodel, freeze-avoiding beetle that overwinters as extremely cold-tolerant larvae in the interior boreal forests of Alaska to temperatures as low as -100 °C. Using a tandem MS-based approach, we compared the proteomes of winter- and summer-collected C.c.p. to identify proteins that may play functional roles in successful overwintering. Using Gene Ontology (GO) analysis and manual interpretation, we identified 104 proteins in winter and 128 proteins in summer samples. We found evidence to indicate a cytoskeletal rearrangement between seasons, with Winter NDSC possessing unique actin and myosin isoforms while summer larvae up-regulated α actinin, tubulin, and tropomyosin. We also detected a fortification of the cuticle in winter via unique cuticle proteins, specifically larval/pupal rigid cuticle protein 66 precursor and larval cuticle protein A2B. Also, of particular interest in the winter larvae was an up-regulation of proteins related to silencing of genes (bromodomain adjacent to zinc finger domain 2A and antisilencing protein 1), proteins involved with metabolism of amines (2-isopropylmalate synthase and dihydrofolate reductase), and immune system process (lysozyme C precursor), among others. This represents the first high throughput MS/MS analysis of a nonmodel, cold-tolerant organism without a concurrent microarray analysis.     

Application of proteomics in biotechnology--microbial proteomics

The genomes of most economically important microbial cells are already sequenced and proteomic technologies can be applied during various process development steps, starting with the selection and optimization of the functions of the industrial strains, application of the knowledge of cell function in response to the changes of production parameters, validation of the downstream processing, and thorough characterization of the final product. Unfortunately, there are only a few direct examples in the literature that present the optimization of the production process based on proteomics. In this review, we discuss the potential of this technology for the design of future bioprocesses and for optimization of existing ones.     

Comprehensive proteomics

Extensive proteome discovery projects using a variety of mass spectrometric techniques have identified proteins matching to 50-70% of the predicted gene models of various species. Comprehensive proteome coverage is desirable for the unbiased comparison of protein quantities between different biological states and for the meaningful comparison of data from multiple samples. Here we discuss the feasibility of this goal in the light of recent technological developments.     

Vectorial proteomics

Vectorial proteomics is a methodology for the differential identification and characterization of proteins and their domains exposed to the opposite sides of biological membranes. Proteomics of membrane vesicles from defined isolated membranes automatically determine cellular localization of the identified proteins and reduce complexity of protein characterizations. The enzymatic shaving of naturally-oriented, or specifically-inverted sealed membrane vesicles, release the surface-exposed peptides from membrane proteins. These soluble peptides are amenable to various chromatographic separations and to sequencing by mass spectrometry, which provides information on the topology of membrane proteins and on their posttranslational modifications. The membrane shaving techniques have made a breakthrough in the identification of in vivo protein phosphorylation sites in membrane proteins form plant photosynthetic and plasma membranes, and from caveolae membrane vesicles of human fat cells. This approach has also allowed investigation of dynamics for in vivo protein phosphorylation in membranes from cells exposed to different conditions. Vectorial proteomics of membrane vesicles with retained peripheral proteins identify extrinsic proteins associated with distinct membrane surfaces, as well as a variety of posttranslational modifications in these proteins. The rapid integration of versatile vectorial proteomics techniques in the functional characterization of biological membranes is anticipated to bring significant insights in cell biology.     

Plastid proteomics

Plastids are essential organelles present in virtually all cells in plants and in green algae. The proteomes of plastids, and in particular of chloroplasts, have received significant amounts of attention in recent years. Various fractionation and mass spectrometry (MS) techniques have been applied to catalogue the chloroplast proteome and its membrane compartments. Neural network and hidden Markov models, in combination with experimentally derived filters, were used to try to predict the chloroplast subproteomes. Some of the many protein-protein interaction, as well as post-translational modifications have been characterized. Nevertheless, our understanding of the chloroplast proteome and its dynamics is very incomplete. Rapid improvements and wide-scale implementation of MS and new tools for comparative proteomics will undoubtedly accelerate this understanding in the near future. Proteomics studies often generate a large amount of data and these data are only meaningful if they can be easily accessed via the 'world-wide-web' and connected to other types of biological information. The plastid proteome data base (PPDB at http://www.ppdb.tc.cornell.edu/) and other web resources are discussed. This review will briefly summarize recent experimental and theoretical efforts, attempt to translate these data into the functions of the chloroplast and outline expectations and possibilities for (comparative) chloroplast proteomics.     

Cancer proteomics

Conclusion The future of cancer diagnostics will be based on a panel of proteomic biomarkers. They could be used to detect cancer at an early stage, to predict and to direct therapies. Enzymes and related proteins are important biological molecules, which could serve as cancer biomarkers. These biomarkers could be intact or fragments of proteins. The challenge is to be able to find and validate these potential biomarkers as clinical diagnostics. With the advances in proteomic technologies, we are closer than ever to find these “new” enzyme molecules or fragments. The translation of newly discovered biomarkers could provide an opportunity to revolutionize the era of personalized medicine.     

Blood-related proteomics

Blood-related proteomics is an emerging field, recently gaining momentum. Indeed, a wealth of data is now available and a plethora of groups has contributed to add pieces to the jigsaw puzzle of protein complexity within plasma and blood cells.In this review article we purported to sail across the mare magnum of the actual knowledge in this research endeavour. The main strides in proteomic investigations on red blood cells, platelets, plasma and white blood cells are hereby presented in a chronological order.Moreover, a glance is given at prospective studies which promise to shift the focus of attention from the end product to its provider, the donor, in a sort of Kantian “Copernican revolution”.A well-rounded portrait of the usefulness of proteomics in blood-related research is accurately given. In particular, proteomic tools could be adopted to follow the main steps of the blood-banking production processes (a comparison of collection methods, pathogen inactivation techniques, storage protocols). Thus proteomics has been recently transformed from a mere basic-research extremely-expensive toy into a dramatically-sensitive and efficient eye-lens to either delve into the depths of the molecular mechanisms of blood and blood components or to establish quality parameters in the blood-banking production chain totally anew.     

Parasite proteomics

Proteomics offers a new set of tools for investigating parasites and parasite-associated disease. In this article, John Barrett, Jim Jefferies and Peter Brophy describe the key technologies involved, including two-dimensional gel electrophoresis, image analysis, biological mass spectroscopy and database searching. The potential applications of proteomics in drug and vaccine discovery are reviewed, as are possible future developments.