Abstract
The microbial degradation of lignocellulose biomass is not only an important biological process but is of increasing industrial significance in the bioenergy sector. The mechanism by which the plant cell wall, an insoluble composite structure, activates the extensive repertoire of microbial hydrolytic enzymes required to catalyze its degradation is poorly understood. Here we have used a transposon mutagenesis strategy to identify a genetic locus, consisting of two genes that modulate the expression of xylan side chain-degrading enzymes in the saprophytic bacterium Cellvibrio japonicus. Significantly, the locus encodes a two-component signaling system, designated AbfS (sensor histidine kinase) and AbfR (response regulator). The AbfR/S two-component system is required to activate the expression of the suite of enzymes that remove the numerous side chains from xylan, but not the xylanases that hydrolyze the beta1,4-linked xylose polymeric backbone of this polysaccharide. Studies on the recombinant sensor domain of AbfS (AbfS(SD)) showed that it bound to decorated xylans and arabinoxylo-oligosaccharides, but not to undecorated xylo-oligosaccharides or other plant structural polysaccharides/oligosaccharides. The crystal structure of AbfS(SD) was determined to a resolution of 2.6A(.) The overall fold of AbfS(SD) is that of a classical Per Arndt Sim domain with a central antiparallel four-stranded beta-sheet flanked by alpha-helices. Our data expand the number of molecules known to bind to the sensor domain of two-component histidine kinases to include complex carbohydrates. The biological rationale for a regulatory system that induces enzymes that remove the side chains of xylan, but not the hydrolases that cleave the backbone of the polysaccharide, is discussed.
Highlights
Saprophytic prokaryotes, such as Cellvibrio japonicus, which utilize the plant cell wall as a major carbon and energy source, synthesize an extensive repertoire of extracellular and membrane-associated hydrolytic enzymes that degrade all the major plant structural polysaccharides, including xylan
Consistent with the complex structure of xylans, the polysaccharide is degraded by a combination of endo-1,4-xylanases [6, 7], referred to as xylanases, and a suite of enzymes that remove the side chains that include ␣-glucuronidases [8], arabinofuranosidases [9, 10], and xylan esterases [11], which remove glucuronic acid, arabinofuranose, and acetyl residues, respectively, from the xylan backbone, whereas the ferulic acidarabinofuranose ester linkage is hydrolyzed by ferulic acid esterases [12]
The complete xylan-degrading system is expressed when C. japonicus is presented with decorated xylans [14], it is unknown whether there is a common inducer for all the enzymes required for complete saccharification of the polysaccharide, or whether different molecules act as inducing triggers for specific subsets of these glycoside hydrolases and esterases
Summary
Saprophytic prokaryotes, such as Cellvibrio japonicus, which utilize the plant cell wall as a major carbon and energy source, synthesize an extensive repertoire of extracellular and membrane-associated hydrolytic enzymes that degrade all the major plant structural polysaccharides, including xylan (for review see Refs. 4, 5). Saprophytic prokaryotes, such as Cellvibrio japonicus, which utilize the plant cell wall as a major carbon and energy source, synthesize an extensive repertoire of extracellular and membrane-associated hydrolytic enzymes that degrade all the major plant structural polysaccharides, including xylan The recent completion of the genome sequence of C. japonicus revealed ϳ150 genes encoding complex carbohydrate-modifying enzymes, primarily glycoside hydrolases and carbohydrate esterases, the majority of which attack the plant cell wall [13]. The complete xylan-degrading system is expressed when C. japonicus is presented with decorated xylans [14], it is unknown whether there is a common inducer for all the enzymes required for complete saccharification of the polysaccharide, or whether different molecules (derived from decorated xylans) act as inducing triggers for specific subsets of these glycoside hydrolases and esterases. The biological significance for a regulatory system that controls the expression of enzymes that remove the side chains but not the xylanases that hydrolyze the backbone of xylans is discussed
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