Mechanisms involved in xyloglucan catabolism by the cellulosome-producing bacterium Ruminiclostridium cellulolyticum.

Julie Ravachol,Pascale De Philip,Romain Borne,Pascal Mansuelle,Stéphanie Perret,Henri-Pierre Fierobe,María J Maté

doi:10.1038/srep22770

Abstract

Xyloglucan, a ubiquitous highly branched plant polysaccharide, was found to be rapidly degraded and metabolized by the cellulosome-producing bacterium Ruminiclostridium cellulolyticum. Our study shows that at least four cellulosomal enzymes displaying either endo- or exoxyloglucanase activities, achieve the extracellular degradation of xyloglucan into 4-glucosyl backbone xyloglucan oligosaccharides. The released oligosaccharides (composed of up to 9 monosaccharides) are subsequently imported by a highly specific ATP-binding cassette transporter (ABC-transporter), the expression of the corresponding genes being strongly induced by xyloglucan. This polysaccharide also triggers the synthesis of cytoplasmic β-galactosidase, α-xylosidase, and β-glucosidase that act sequentially to convert the imported oligosaccharides into galactose, xylose, glucose and unexpectedly cellobiose. Thus R. cellulolyticum has developed an energy-saving strategy to metabolize this hemicellulosic polysaccharide that relies on the action of the extracellular cellulosomes, a highly specialized ABC-transporter, and cytoplasmic enzymes acting in a specific order. This strategy appears to be widespread among cellulosome-producing mesophilic bacteria which display highly similar gene clusters encoding the cytosolic enzymes and the ABC-transporter.

Highlights

Xyloglucan, a ubiquitous highly branched plant polysaccharide, was found to be rapidly degraded and metabolized by the cellulosome-producing bacterium Ruminiclostridium cellulolyticum
The polysaccharide was completely converted into a mixture of 4-glucosyl xyloglucan oligosaccharides (XGO4) XXXG, XLXG, XXLG and XLLG (Fig. 1) as shown by analysis using High Pressure Anion Exchange Chromatography coupled with Pulsed Amperometric Detection (HPAEC-PAD)
R. cellulolyticum was found to grow on xyloglucan oligosaccharides but displayed a slightly extended doubling time (17 ± 0.4 h) compared to that determined on xyloglucan-based medium, that might be due to a carbon overflow of the glycolytic pathway[29,30]

Summary

Introduction

Xyloglucan, a ubiquitous highly branched plant polysaccharide, was found to be rapidly degraded and metabolized by the cellulosome-producing bacterium Ruminiclostridium cellulolyticum. One of the most abundant hemicellulosic polymers is xylan, whose backbone is composed of β -1,4 linked xylosyl residues, which can display a number of side chains which vary tremendously among plants, or among tissues within the same plant[1] Xyloglucan is another prominent hemicellulosic polysaccharide, especially in primary cell walls[2,3], where it interacts with the cellulose microfibrils. Most anaerobic cellulolytic bacteria gather the plant cell wall degrading enzymes in million-Da range extracellular complexes termed cellulosomes[6,7,8,9], sparingly produced compared to aerobic microorganisms The simplest cellulosomes such as those produced by the model bacterium Ruminiclostridium cellulolyticum (formerly known as Clostridium cellulolyticum10) are composed of a single primary scaffoldin which binds to crystalline cellulose via a family-3a Carbohydrate Binding Module (CBM)[11]. The cellulosomes synthesized by other anaerobic bacteria, such as R. thermocellum (formerly known as Clostridium thermocellum10) or Ruminococcus flavefaciens, can be much more complex and include several interacting scaffoldins, as well as several types of specific cohesin/dockerin docking devices[9]

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Results

Conclusion