Abstract
BackgroundExpansins are relatively small proteins that lack enzymatic activity and are found in plants and microorganisms. The function of these proteins is to disrupt the plant cell walls by interfering with the non-covalent interchain bonding of the polysaccharides. Expansins were found to be important for plant growth, but they are also expressed by various bacteria known to have interactions with plants. Clostridium clariflavum is a plant cell wall-degrading bacterium with a highly elaborate cellulosomal system. Among its numerous dockerin-containing genes, two expansin-like proteins, Clocl_1862 and Clocl_1298 (termed herein CclEXL1 and CclEXL2) were identified, and CclEXL1 was found to be expressed as part of the cellulosome system. This is the first time that an expansin-like protein is identified in a cellulosome complex, which implicates its possible role in biomass deconstruction.ResultsIn the present article, we analyzed the functionality of CclEXL1. Its dockerin was characterized and shown to bind selectively to type-I cohesins of C. clariflavum, with preferential binding to the cohesin of ScaG, and additionally to a type-I cohesin of C. cellulolyticum. We demonstrated experimentally that the expansin-like protein binds preferentially to microcrystalline cellulose, but it also binds to acid-swollen cellulose, xylan, and wheat straw. CclEXL1 exhibited a pronounced loosening effect on filter paper, which resulted in substantial decrease in tensile stress. The C. clariflavum expansin-like protein thus enhances significantly enzymatic hydrolysis of cellulose, both by C. clariflavum cellulosomes and two major cellulosomal cellulases from this bacterium: GH48 (exoglucanase) and GH9 (endoglucanase). Finally, we demonstrated CclEXL1-mediated enhancement of microcrystalline cellulose degradation by different cellulosome fractions and the two enzymes.ConclusionsThe results of this study confirm that the C. clariflavum expansin-like protein is part of the elaborate cellulosome system of this bacterium with capabilities of cellulose creeping. The data suggest that pretreatment of cellulosic materials with CclEXL1 can bring about substantial improvement of hydrolysis by cellulases.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-016-0474-5) contains supplementary material, which is available to authorized users.
Highlights
Expansins are relatively small proteins that lack enzymatic activity and are found in plants and microorganisms
CclEXL1 was only identified in the lower-molecular-weight cellulosome fractions, and at relatively low expression levels (2 % relative to the amount of primary scaffoldin, ScaA, in this fraction)
We isolated the cellulosomes of C. clariflavum, separated between the high-molecularweight (MCCI) and the low-molecular-weight (MCCII) complexes by gel filtration, and we examined whether addition of CclEXL1 would result in a synergistic effect on cellulose degradation (Fig. 5)
Summary
Expansins are relatively small proteins that lack enzymatic activity and are found in plants and microorganisms. Among its numerous dockerin-containing genes, two expansin-like proteins, Clocl_1862 and Clocl_1298 (termed CclEXL1 and CclEXL2) were identified, and CclEXL1 was found to be expressed as part of the cellulosome system. This is the first time that an expansin-like protein is identified in a cellulosome complex, which implicates its possible role in biomass deconstruction. Major efforts have been invested in recent years to find alternative fuel sources, and one of these is the use of lignocellulosic biomass for production of bioethanol [1,2,3] This initiative has led to a growing interest in cellulolytic microorganisms and their.
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