Bacterial protein meta-interactomes predict cross-species interactions and protein function.

BACKGROUND: Protein-protein interactions (PPIs) can offer compelling evidence for protein function, especially when viewed in the context of proteome-wide interactomes. Bacteria have been popular subjects of interactome studies: more than six different bacterial species have been the subjects of comprehensive interactome studies while several more have had substantial segments of their proteomes screened for interactions. The protein interactomes of several bacterial species have been completed, including several from prominent human pathogens. The availability of interactome data has brought challenges, as these large data sets are difficult to compare across species, limiting their usefulness for broad studies of microbial genetics and evolution. RESULTS: In this study, we use more than 52,000 unique protein-protein interactions (PPIs) across 349 different bacterial species and strains to determine their conservation across data sets and taxonomic groups. When proteins are collapsed into orthologous groups (OGs) the resulting meta-interactome still includes more than 43,000 interactions, about 14,000 of which involve proteins of unknown function. While conserved interactions provide support for protein function in their respective species data, we found only 429 PPIs (~1% of the available data) conserved in two or more species, rendering any cross-species interactome comparison immediately useful. The meta-interactome serves as a model for predicting interactions, protein functions, and even full interactome sizes for species with limited to no experimentally observed PPI, including Bacillus subtilis and Salmonella enterica which are predicted to have up to 18,000 and 31,000 PPIs, respectively. CONCLUSIONS: In the course of this work, we have assembled cross-species interactome comparisons that will allow interactomics researchers to anticipate the structures of yet-unexplored microbial interactomes and to focus on well-conserved yet uncharacterized interactors for further study. Such conserved interactions should provide evidence for important but yet-uncharacterized aspects of bacterial physiology and may provide targets for anti-microbial therapies.

Proteomic and functional analysis of the cellulase system expressed by Postia placenta during brown rot of solid wood.

Brown rot basidiomycetes have an important ecological role in lignocellulose recycling and are notable for their rapid degradation of wood polymers via oxidative and hydrolytic mechanisms. However, most of these fungi apparently lack processive (exo-acting) cellulases, such as cellobiohydrolases, which are generally required for efficient cellulolysis. The recent sequencing of the Postia placenta genome now permits a proteomic approach to this longstanding conundrum. We grew P. placenta on solid aspen wood, extracted proteins from the biodegrading substrate, and analyzed tryptic digests by shotgun liquid chromatography-tandem mass spectrometry. Comparison of the data with the predicted P. placenta proteome revealed the presence of 34 likely glycoside hydrolases, but only four of these--two in glycoside hydrolase family 5, one in family 10, and one in family 12--have sequences that suggested possible activity on cellulose. We expressed these enzymes heterologously and determined that they all exhibited endoglucanase activity on phosphoric acid-swollen cellulose. They also slowly hydrolyzed filter paper, a more crystalline substrate, but the soluble/insoluble reducing sugar ratios they produced classify them as nonprocessive. Computer simulations indicated that these enzymes produced soluble/insoluble ratios on reduced phosphoric acid-swollen cellulose that were higher than expected for random hydrolysis, which suggests that they could possess limited exo activity, but they are at best 10-fold less processive than cellobiohydrolases. It appears likely that P. placenta employs a combination of oxidative mechanisms and endo-acting cellulases to degrade cellulose efficiently in the absence of a significant processive component.

Differential expression in Phanerochaete chrysosporium of membrane-associated proteins relevant to lignin degradation.

Fungal lignin-degrading systems likely include membrane-associated proteins that participate in diverse processes such as uptake and oxidation of lignin fragments, production of ligninolytic secondary metabolites, and defense of the mycelium against ligninolytic oxidants. Little is known about the nature or regulation of these membrane-associated components. We grew the white rot basidiomycete Phanerochaete chrysosporium on cellulose or glucose as the carbon source and monitored the mineralization of a (14)C-labeled synthetic lignin by these cultures to assess their ligninolytic competence. The results showed that the cellulose-grown cultures were ligninolytic, whereas the glucose-grown ones were not. We isolated microsomal membrane fractions from both types of culture and analyzed tryptic digests of their proteins by shotgun liquid chromatography-tandem mass spectrometry. Comparison of the results against the predicted P. chrysosporium proteome showed that a catalase (Joint Genome Institute P. chrysosporium protein identification number [I.D.] 124398), an alcohol oxidase (126879), two transporters (137220 and 132234), and two cytochrome P450s (5011 and 8912) were upregulated under ligninolytic conditions. Quantitative reverse transcription-PCR assays showed that RNA transcripts encoding all of these proteins were also more abundant in ligninolytic cultures. Catalase 124398, alcohol oxidase 126879, and transporter 137220 were found in a proteomic analysis of partially purified plasma membranes from ligninolytic P. chrysosporium and are therefore most likely associated with the outer envelope of the fungus.

New insights into the ligninolytic capability of a wood decay ascomycete.

Wood-grown cultures of Daldinia concentrica oxidized a permethylated beta-(14)C-labeled synthetic lignin to (14)CO(2) and also cleaved a permethylated alpha-(13)C-labeled synthetic lignin to give C(alpha)-C(beta) cleavage products that were detected by (13)C nuclear magnetic resonance spectrometry. Therefore, this ascomycete resembles white-rot basidiomycetes in attacking the recalcitrant nonphenolic structures that predominate in lignin.