Dataset Information

Spatial constraints drive amylosome-mediated resistant starch degradation by Ruminococcus bromii in the human colon

ABSTRACT: Degradation of complex dietary fibers by gut microbes is essential for colonic fermentation, short-chain fatty acid production, and microbiome function. Ruminococcus bromii is the primary resistant starch (RS) degrader in humans, which relies on the amylosome, a specialized cell-bound enzymatic complex. To unravel its structure-function relationship and the interplay among its components, we applied an holistic multilayered approach and found that amylosome combinatorics, resistant starch degradation and enzymatic synergy are regulated at two levels: structural constraints enforcing enzyme proximity and expression-driven shifts in enzyme proportions. Cryo-electron tomography revealed that the amylosome comprises a constitutive extracellular layer extending toward the RS. However, proteomics demonstrated its remodeling across different growth conditions, with Amy4 and Amy16 comprising 60% of the amylosome in response to RS. Structural and biochemical analyses revealed complementarity and synergistic RS degradation by these enzymes, which allow R. bromii to fine-tune its adaptation to dietary fiber and shape colonic metabolism

INSTRUMENT(S):

ORGANISM(S): Ruminococcus Bromii L2-63

SUBMITTER: Anke Trautwein-Schult

LAB HEAD: Professor Dr.Dörte

PROVIDER: PXD060149 | Pride | 2025-11-02

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	201211_O1_P7_SG_ATS_F10_1.raw	Raw
	201211_O1_P7_SG_ATS_F10_2.raw	Raw
	201211_O1_P7_SG_ATS_F10_3.raw	Raw
	201211_O1_P7_SG_ATS_F14_1.raw	Raw
	201211_O1_P7_SG_ATS_F14_2.raw	Raw

Items per page:

1 - 5 of 225

Similar Datasets

Project description:The analysis of fungi secreted proteins has been increasingly employed as a powerful strategy in the prospecting of new catalysts with potential biotechnological application. Since enzyme production is strongly modulated by several factors such as pH, available nutrients, water content, oxygen levels and temperature, the evaluation of growth conditions is of utmost importance to achieve optimal enzyme production. Here, a non-sequenced wood-rotting fungus, L. crinitus, was selected as a model organism for secretome analysis by means of in vitro enzymatic assays and in-depth characterization via proteomics. For enzyme production, the fungus was cultured with several types and concentrations of carbon and nitrogen sources and variable water content. Five different carbon substrates (glucose, maltose, starch, sucrose, carboxymethylcellulose (CMC), glycerol and fructose) and three nitrogen containing compounds (urea, sodium nitrate and ammonium chloride) were used as substitutes of the original carbon and nitrogen sources in culture media. Interestingly, the biomass yield as well as the array of secreted proteins differed drastically under different growing conditions. A mixture of soluble secreted extracts derived from different culture conditions was analyzed both by shotgun mass spectrometry and through protein separation by two-dimensional gel electrophoresis (2DE) prior to identification via LC-MS/MS. Proteins were identified by sequence homology searches using mass spectrometry (MS)-driven BLAST. The spectrum of secreted enzymes comprised various types of CAZymes (carbohydrate-active enzymes), oxidase/reductases, proteases, lipase/esterases, proteins with non-related functions thereby classified as miscellany proteins and hypothetical or unknown predicted proteins. Although pre-separation by 2DE improved the number of identifications (protein map of 150 spots corresponding to 171 identifications) compared to the shotgun approach (98 identifications) both strategies revealed similar distribution of proteins within the functional categories described above. Culture media with reduced water content stimulated the expression of oxidases/reductases such as Lac, MnP, GMC oxidoreductases and glyoxal oxidase while hydrolases were induced during submerged fermentation. The diversity of proteins observed within both the CAZyme and oxidoreductase groups revealed in this fungus a powerful arsenal of enzymes dedicated to the breakdown and consumption of lignocellulose. Moreover, further secretome characterization after sequencing and analysis of the L. crinitus genome can potentially lead to the discovery of novel enzymes of industrial and biotechnological interest. Importantly, information on sequencing data could also reveal novel enzymes which production is strongly stimulated only in specific growing conditions.

Dataset Information

Spatial constraints drive amylosome-mediated resistant starch degradation by Ruminococcus bromii in the human colon

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets