Lignin is a heterogeneous aromatic biopolymer that accounts for nearly 30% of the organic carbon on Earth and is one of the few renewable sources of aromatic chemicals. As the most recalcitrant of ...the three components of lignocellulosic biomass (cellulose, hemicellulose and lignin), lignin has been treated as a waste product in the pulp and paper industry, where it is burned to supply energy and recover pulping chemicals in the operation of paper mills. Extraction of higher value from lignin is increasingly recognized as being crucial to the economic viability of integrated biorefineries. Depolymerization is an important starting point for many lignin valorization strategies, because it could generate valuable aromatic chemicals and/or provide a source of low-molecular-mass feedstocks suitable for downstream processing. Commercial precedents show that certain types of lignin (lignosulphonates) may be converted into vanillin and other marketable products, but new technologies are needed to enhance the lignin value chain. The complex, irregular structure of lignin complicates chemical conversion efforts, and known depolymerization methods typically afford ill-defined products in low yields (that is, less than 10-20wt%). Here we describe a method for the depolymerization of oxidized lignin under mild conditions in aqueous formic acid that results in more than 60wt% yield of low-molecular-mass aromatics. We present the discovery of this facile C-O cleavage method, its application to aspen lignin depolymerization, and mechanistic insights into the reaction. The broader implications of these results for lignin conversion and biomass refining are also considered.
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Dostopno za:
DOBA, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Calorie restriction (CR) extends life span in diverse species. Mitochondria play a key role in CR adaptation; however, the molecular details remain elusive. We developed and applied a quantitative ...mass spectrometry method to probe the liver mitochondrial acetyl-proteome during CR versus control diet in mice that were wild-type or lacked the protein deacetylase SIRT3. Quantification of 3,285 acetylation sites—2,193 from mitochondrial proteins—rendered a comprehensive atlas of the acetyl-proteome and enabled global site-specific, relative acetyl occupancy measurements between all four experimental conditions. Bioinformatic and biochemical analyses provided additional support for the effects of specific acetylation on mitochondrial protein function. Our results (1) reveal widespread reprogramming of mitochondrial protein acetylation in response to CR and SIRT3, (2) identify three biochemically distinct classes of acetylation sites, and (3) provide evidence that SIRT3 is a prominent regulator in CR adaptation by coordinately deacetylating proteins involved in diverse pathways of metabolism and mitochondrial maintenance.
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► MS quantifies 1,578 mitochondrial acetyl sites altered during CR and loss of SIRT3 ► SIRT3 functions as a prominent regulator in CR adaptation ► CR and SIRT3 regulate previously unrecognized processes in mitochondria ► We provide an acetylation atlas for understanding mitochondrial regulation in CR
Protein glycosylation is a highly important, yet poorly understood protein post-translational modification. Thousands of possible glycan structures and compositions create potential for tremendous ...site heterogeneity. A lack of suitable analytical methods for large-scale analyses of intact glycopeptides has limited our abilities both to address the degree of heterogeneity across the glycoproteome and to understand how this contributes biologically to complex systems. Here we show that N-glycoproteome site-specific microheterogeneity can be captured via large-scale glycopeptide profiling methods enabled by activated ion electron transfer dissociation (AI-ETD), ultimately characterizing 1,545 N-glycosites (>5,600 unique N-glycopeptides) from mouse brain tissue. Our data reveal that N-glycosylation profiles can differ between subcellular regions and structural domains and that N-glycosite heterogeneity manifests in several different forms, including dramatic differences in glycosites on the same protein. Moreover, we use this large-scale glycoproteomic dataset to develop several visualizations that will prove useful for analyzing intact glycopeptides in future studies.
How dissociation is effected determines the upstream sample handling whereas the spectral features it produces regulate the downstream informatics approach. (To listen to a podcast about this ...feature, please go to the Analytical Chemistry website at pubs.acs.org/journal/ancham.
The existence of extracellular phosphoproteins has been acknowledged for over a century. However, research in this area has been undeveloped largely because the kinases that phosphorylate secreted ...proteins have escaped identification. Fam20C is a kinase that phosphorylates S-x-E/pS motifs on proteins in milk and in the extracellular matrix of bones and teeth. Here, we show that Fam20C generates the majority of the extracellular phosphoproteome. Using CRISPR/Cas9 genome editing, mass spectrometry, and biochemistry, we identify more than 100 secreted phosphoproteins as genuine Fam20C substrates. Further, we show that Fam20C exhibits broader substrate specificity than previously appreciated. Functional annotations of Fam20C substrates suggest roles for the kinase beyond biomineralization, including lipid homeostasis, wound healing, and cell migration and adhesion. Our results establish Fam20C as the major secretory pathway protein kinase and serve as a foundation for new areas of investigation into the role of secreted protein phosphorylation in human biology and disease.
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•Fam20C is unique among the known secretory pathway kinases•Fam20C generates the majority of the secreted phosphoproteome•Fam20C substrates are implicated in a broad spectrum of biological processes•Fam20C is crucial for proper adhesion, migration, and invasion of breast cancer cells
The kinases that catalyze the phosphorylation of secreted proteins have only recently been identified, with Fam20C being identified as the kinase responsible for generating the vast majority of the secreted phosphoproteome, including substrates thought to drive tumor cell migration.
Selected reaction monitoring on a triple quadrupole mass spectrometer is currently experiencing a renaissance within the proteomics community for its, as yet, unparalleled ability to characterize and ...quantify a set of proteins reproducibly, completely, and with high sensitivity. Given the immense benefit that high resolution and accurate mass instruments have brought to the discovery proteomics field, we wondered if highly accurate mass measurement capabilities could be leveraged to provide benefits in the targeted proteomics domain as well. Here, we propose a new targeted proteomics paradigm centered on the use of next generation, quadrupole-equipped high resolution and accurate mass instruments: parallel reaction monitoring (PRM). In PRM, the third quadrupole of a triple quadrupole is substituted with a high resolution and accurate mass mass analyzer to permit the parallel detection of all target product ions in one, concerted high resolution mass analysis. We detail the analytical performance of the PRM method, using a quadrupole-equipped bench-top Orbitrap MS, and draw a performance comparison to selected reaction monitoring in terms of run-to-run reproducibility, dynamic range, and measurement accuracy. In addition to requiring minimal upfront method development and facilitating automated data analysis, PRM yielded quantitative data over a wider dynamic range than selected reaction monitoring in the presence of a yeast background matrix because of PRM's high selectivity in the mass-to-charge domain. With achievable linearity over the quantifiable dynamic range found to be statistically equal between the two methods, our investigation suggests that PRM will be a promising new addition to the quantitative proteomics toolbox.
Mitochondria are essential for numerous cellular processes, yet hundreds of their proteins lack robust functional annotation. To reveal functions for these proteins (termed MXPs), we assessed ...condition-specific protein-protein interactions for 50 select MXPs using affinity enrichment mass spectrometry. Our data connect MXPs to diverse mitochondrial processes, including multiple aspects of respiratory chain function. Building upon these observations, we validated C17orf89 as a complex I (CI) assembly factor. Disruption of C17orf89 markedly reduced CI activity, and its depletion is found in an unresolved case of CI deficiency. We likewise discovered that LYRM5 interacts with and deflavinates the electron-transferring flavoprotein that shuttles electrons to coenzyme Q (CoQ). Finally, we identified a dynamic human CoQ biosynthetic complex involving multiple MXPs whose topology we map using purified components. Collectively, our data lend mechanistic insight into respiratory chain-related activities and prioritize hundreds of additional interactions for further exploration of mitochondrial protein function.
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•PPI mapping of 50 MXPs reveals mitochondrial protein functions•C17orf89 is a CI assembly factor depleted in a case of CI deficiency•LYRM5 interacts with and deflavinates the electron transferring flavoprotein•Proteins involved in coenzyme Q biosynthesis form a dynamic “complex Q”
Mitochondria are essential organelles, yet hundreds of their proteins lack robust functional characterization. Floyd et al. (2016) define interaction partners for 50 such proteins, providing hypotheses about their roles in mitochondria. In particular, their work lends mechanistic insight into respiratory chain activities related to complex I, the electron transferring flavoprotein, and coenzyme Q.
From plant research to biomedicine, proteome analysis plays a critical role in many areas of biological inquiry. Steady improvement in mass spectrometer (MS) technology has transformed the speed and ...depth of proteome analysis. Proteomes of simple organisms can now be sequenced to near completion in just over an hour. Comparable coverage of mammalian proteomes, however, still requires hours or even days of analysis. Here we ask why current technology fails to achieve comprehensive and rapid analysis of the more complex mammalian proteomes. We propose that further advancements in MS technology alone are unlikely to solve this problem and suggest that concomitant improvements in peptide separation technology will be critical.
From plant research to biomedicine, proteome analysis plays a critical role in many areas of biological inquiry. Steady improvement in mass spectrometer (MS) technology has transformed the speed and depth of proteome analysis. Proteomes of simple organisms can now be sequenced to near completion in just over an hour. Comparable coverage of mammalian proteomes, however, still requires hours or even days of analysis. Here we ask why current technology fails to achieve comprehensive and rapid analysis of the more complex mammalian proteomes. We propose that further advancements in MS technology alone are unlikely to solve this problem and suggest that concomitant improvements in peptide separation technology will be critical.
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