Combined Crystal Structure of a Type I Cohesin: Mutation and Affinity Binding Studies Reveal Structural Determinants of Cohesin-Dockerin Specificities

Abstract Cohesin-dockerin interactions orchestrate the assembly of one of nature's most elaborate multi-enzyme complexes, the cellulosome. Cellulosomes are produced exclusively by anaerobic microbes and mediate highly efficient hydrolysis of plant structural polysaccharides, such as cellulose and hemicellulose. In the canonical model of cellulosome assembly, type-I dockerin modules of the enzymes bind to reiterated type-I cohesin modules of a primary scaffoldin. Each type-I dockerin contains two highly conserved cohesin-binding sites, which confer quaternary flexibility to the multienzyme complex. The scaffoldin also bears a type-II dockerin that anchors the entire complex to the cell surface by binding type-II cohesins of anchoring scaffoldins. In Bacteroides cellulosolvens, however, the organisation of the cohesin-dockerin types is reversed, whereby type-II cohesin-dockerin pairs integrate the enzymes into the primary scaffoldin and type-I modules mediate cellulosome attachment to an anchoring scaffoldin. Here we report the crystal structure of a type-I cohesin from B. cellulosolvens anchoring scaffoldin ScaB, to 1.84-A resolution. The structure resembles other type-I cohesins, and the putative dockerin-binding site, centred at beta;-strands 3, 5 and 6, is likely to be conserved in other B. cellulosolvens type-I cohesins. Combined computational modelling, mutagenesis and affinity-based binding studies revealed similar hydrogen-bonding networks between putative Ser/Asp recognition residues, in the dockerin at positions 11/12 and 45/46, suggesting that a dual-binding mode is not exclusive to the integration of enzymes into primary cellulosomes but can also characterise polycellulosome assembly and cell-surface attachment. This general approach may provide valuable structural information of the cohesin-dockerin interface, in lieu of a definitive crystal structure.