Proteomic Insights into the Cellulose Degradation Systems of Cytophaga hutchinsonii and Sporocytophaga myxococcoides

Cytophaga hutchinsonii and Sporocytophaga myxococcoides are both Gram-negative, aerobic, mesophilic, cellulose degrading bacterium belonging to the phylum Bacteroidetes.\ua0 Despite both of these organisms being isolated for almost a century, they remain poorly studied with the proteins responsible for cellulose degradation remaining unknown.\ua0 Both organisms utilize cell associated machinery in order to degrade crystalline cellulose but do not contain any expected scaffoldin or dockerin proteins of the well-known complexed cellulosome system.\ua0 Therefore, we have employed proteomic analysis in order to identify the abundant proteins present during growth on crystalline cellulose and assess their putative roles in cellulose hydrolysis.\ua0 Further, we have been able to infer the localization of these proteins based on their abundance in specific cellular fractions.\ua0 Taken together, the abundance and localization of proteins putatively involved in cellulose degradation led into important insights into the cellulose degradation mechanisms in both C. hutchinsonii and S. myxococcoides

Mapping the Enzyme Machineries of Cellulolytic Soil-Dwelling Bacteroidetes

Bacteria from the phylum Bacteroidetes are regarded as proficient degraders of complex carbohydrates, but most species are limited to soluble glycans, e.g. hemicelluloses and pectins. Two aerobic Bacteroidetes members, Cytophaga hutchinsonii and Sporocytophaga myxococcoides, have however been known as proficient cellulose metabolizers for decades, but do not conform to the known mechanisms of enzymatic cellulose conversion. Neither species encodes cellobiohydrolases or lytic polysaccharide monooxygenases, and no apparent\ua0\ua0 complexed systems such as cellulosomes have been identified.\ua0\ua0 Many Bacteroidetes species utilize so-called polysaccharide utilization loci (PULs) which encode the necessary enzymes, binding proteins, sugar transporters and regulatory elements for target polysaccharides, but also these are absent in the genomes of C. hutchinsonii and S. myxococcoides. Mutagenesis studies instead point toward the Type IX secretion system being a crucial factor in polysaccharide turnover, and it is also tightly linked to their rapid gliding motility. In order to shed light on the enigmatic cellulolytic systems of these bacteria, we have used quantitative proteomics to map which proteins they produce during growth on cellulose and pectin, respectively, and determined the proteins’ cellular locations. Both bacteria produced similar yet distinct arrays of mostly unstudied putative cellulases during growth, and interestingly, cellulolytic activity was detected not only in the extracellular fraction and outer membrane but also intracellularly. In addition, several glycoside hydrolase family 8 (GH8) enzymes, that have previously been overlooked as potential cellulases in these species, were found to be both abundant and selectively produced during growth on cellulose. These GH8-containing proteins, which comprise large regions of unknown function and range between ~1100-2800 amino acids in total, are currently being functionally characterized to clarify their roles in cellulose turnover

Reduced catabolic protein expression in Clostridium butyricum DSM 10702 correlate with reduced 1,3-propanediol synthesis at high glycerol loading

WOS: 000358064700001PubMed ID: 25401066Higher initial glycerol loadings (620 mM) have a negative effect on growth and 1,3-propanediol (1,3-PDO) synthesis in Clostridium butyricum DSM 10702 relative to lower initial glycerol concentrations (170 mM). To help understand metabolic shifts associated with elevated glycerol, protein expression levels were quantified by LC/MS/MS analyses. Thirty one (31) proteins involved in conversion of glycerol to 1,3-PDO and other by-products were analyzed by multiple reaction monitoring (MRM). The analyses revealed that high glycerol concentrations reduced cell growth. The expression levels of most proteins in glycerol catabolism pathways were down-regulated, consistent with the slower growth rates observed. However, at high initial glycerol concentrations, some of the proteins involved in the butyrate synthesis pathways such as a putative ethanol dehydrogenase (CBY_3753) and a 3-hydroxybutyryl-CoA dehydrogenase (CBY_3045) were up-regulated in both exponential and stationary growth phases. Expression levels of proteins (CBY_0500, CBY_0501 and CBY_0502) involved in the reductive pathway of glycerol to 1,3-PDO were consistent with glycerol consumption and product concentrations observed during fermentation at both glycerol concentrations, and the molar yields of 1,3-PDO were similar in both cultures. This is the first report that correlates expression levels of glycerol catabolism enzymes with synthesis of 1,3-PDO in C. butyricum. The results revealed that significant differences in the expression of a small subset of proteins were observed between exponential and stationary growth phases at both low and high glycerol concentrations.Scientific And Technological Research Council Of Turkey (TUBITAK)Turkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [BIDEB 2214]; Scientific And Technological Research Council Of Turkey (CAYDAG Project)Turkiye Bilimsel ve Teknolojik Arastirma Kurumu (TUBITAK) [109Y150]; Genome CanadaGenome Canada; Province of ManitobaThe authors would like to acknowledge Prof. Dr. Nuri Azbar for his valuable contribution and insightful discussions. The authors would like to thank Dr. Graham Alvare for his technical assistance with bioinformatics. This work was funded by The Scientific And Technological Research Council Of Turkey (TUBITAK, BIDEB 2214 and CAYDAG Project No: 109Y150), by Genome Canada, through the Applied Genomics Research in Bioproducts or Crops (ABC) program for the grant titled, "Microbial Genomics for Biofuels and CoProducts from Biorefining Processes", and by the Province of Manitoba, through the Manitoba Research Innovation Fund (MRIF)

Economic Diplomacy of China in Africa

This study is about economic diplomacy of China in Africa. It describes development of China-Africa relations, their current form and examines, whether less democratic countries trade more with China than the democratic ones. The study is divided into four chapters. The first one defines economic diplomacy theoretically. The second one focuses on China-Africa relationship from its history up to establishing of Forum on China-Africa Cooperation. It also describes motives of China-Africa cooperation, stressing China's energy dependance. The third chapter describes development of China-Africa trade, Chinese foreign direct investments and development aid and pros and cons of China's presence in Africa. The last chapter examines relationship between level of democracy in african countries and their trade with China

Additional file 6: of Transcriptomic and proteomic analyses of core metabolism in Clostridium termitidis CT1112 during growth on α-cellulose, xylan, cellobiose and xylose

Expression values and Znet values of genes and gene products identified in C. termitidis cultured under various substrate conditions. Normalized proteomic Z-scores Pnet are represented as: (i) P0: xylose grown cells – cellobiose grown cells; (ii) P1: α-cellulose grown cells – cellobiose grown cells; and (iii) P2: xylan grown cells – cellobiose grown cells. Normalized transcriptomic Z-scores Rnet are represented as: (i) R0: xylose grown cells – cellobiose grown cells; (ii) R1: α-cellulose grown cells – cellobiose grown cells; and (iii) R2: xylan grown cells – cellobiose grown cells. For any protein or RNA transcripts, a negative value represents higher expression on cellobiose, while a positive value represents higher expression in any of the other corresponding substrate. Signal to Noise ratios (S/N) of individual genes under two comparing conditions are given. S/N0: xylose grown cells vs cellobiose grown cells; S/N1: α-cellulose grown cells – cellobiose grown cells; S/N2: xylan grown cells – cellobiose grown cells. Genes with a S/N value 2.8 or greater have a false discovery rate (FDR) 10 % or less. CB, Cellobiose; AC, α-cellulose; Numbers in brackets indicate biological replicate. (XLSX 817 kb