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Johan 12May2014

Johan 12May2014. Proposed model of activation of σ I factors. I ntroduction. Cellulosome-producing bacterium Contains a variety of carbohydrate active enzymes in order to break down complex carbohydrates

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Johan 12May2014

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  1. Johan 12May2014

  2. Proposed model of activation of σI factors

  3. Introduction • Cellulosome-producingbacterium • Contains a variety of carbohydrateactiveenzymes in order to break down complex carbohydrates • Divers nature of biomassrequires a regulated mixture of degradingenzymes • Regulationcellulosomalgenes

  4. Scematicrepresentaion domain organization of RsgI-likeproteins in C. thermocellum

  5. Overall structuralfeatures of the threestructures RsgI-CBMsexhibitstructuralcharacteristicstypical of family 3 CBMs: Nine-stranded antiparallel immunoglobulin –likebeta –sandwich Ca is often found in carbohydrate- recognitiondomains but in this case the Ca is to far awayfromsugar binding surface In these structures, Ca has a structuralrole

  6. C-termini form a network of interactionsthat anchor the last residues of the proteinsto the beta-sanswichfold. Last 5 residues are nearlyconserved. Last aa is a prolinewhich is sandwichedbetweentwotyrosinerings

  7. Carbohydrate binding RsgI-CBM3s show different carbohydrate binding preferences, most likelyowingtodiffenrences in the lengthsandamino-acid compositions of theircarbohydrate binding surfaces 0059CBM3b: Broad specificity pattern (crystalline, amorphous cellulose); bottom beta sheet comprising strands 1-2-4-7 is flat (green fig 11a) and contains a long array of comprising 4 aromatic residues. 0267CBM3b: bindstobothcrystallineandamorphous cellulose, same lengt binding stip as 0404CBM3b but containsan extra loop 0404CBM3b: weaksignalforxyloglucan (data notshown) dueto short carbohydrate binding stip

  8. 0059CBM3b (green) 0267CBM3b (red) 0404CBM3b (blue)

  9. Cellulose binding for 0267CBM variants Alanine mutation screening shows that the residues in the loop are significatfor cellulose binding The position of the loop (at the edge of the binding planewouldallowitto fit along the edge of cellulose crystals.

  10. Summary Recently a putativecarbohydrate- biosensing system was descovered Extra cellularprobelikeproteinsdetectpolysaccharidecomponentsandsubsequently transfer a signalto the cell, activatingtranscription of target genes. Three of those family 3 CBMs have been shownto sense the presence of bothcrystallineandamorphous cellulose, xylanxyloglucanandgalactan. Their 3D stucturesweredetermined in thiswork The presense of numerousRsgIproteinswithunique c-terninalCMBssugeststhateach of themcouldberesponsiblefor the regulation of a specific set of genes Therefore, the biomasssensinganddeconstructionmachinerymathes the dynamicnature of the biomass.

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