A novel decrystallizing protein CxEXL22 from Arthrobotrys sp. CX1 capable of synergistically hydrolyzing cellulose with cellulases

Abstract

A novel expansin-like protein (CxEXL22) has been identified and characterized from newly isolated Arthrobotrys sp. CX1 that can cause cellulose decrystallization. Unlike previously reported expansin-like proteins from microbes, CxEXL22 has a parallel β-sheet domain at the N terminal, containing many hydrophobic residues to form the hydrophobic surface as part of the groove. The direct phylogenetic relationship implied the genetic transfers occurred from nematode to nematicidal fungal Arthrobotrys sp. CX1. CxEXL22 showed strong activity for the hydrolysis of hydrogen bonds between cellulose molecules, especially when highly crystalline cellulose was used as substrate. The hydrolysis efficiency of Avicel was increased 7.9-fold after pretreating with CxEXL22. The rupture characterization of crystalline region indicated that CxEXL22 strongly binds cellulose and breaks up hydrogen bonds in the crystalline regions of cellulose to split cellulose chains, causing significant depolymerization to expose much more microfibrils and enhances cellulose accessibility.

Pretreatment of chitin depolymerization by expansin-like protein to improve hydrolysis efficiency

“…This finding is consistent with a previous study on the synergism of expansin-like proteins [ 29 ]. As previously reported, incubation of chitin with CxEXL22 or HcEXL alone did not result in significant production of reducing sugars [ 17 , 18 ]. Compared to non-pretreated chitin, chitinase activity increased by 2- and 3.4-fold, respectively, after chitin was pretreated with CxEXL22 or HcEXL for 14 h ( Fig.…”

“…CxEXL22 and HcEXL have previously been reported to disrupt hydrogen bonding between matrix polysaccharides and cellulose microfibrils, causing them to swell [ 17 , 18 ]. After expansin-like protein pretreatment, the crystallinity index of chitin decreased and the d-spacing increased slightly owing to swelling.…”

“…CxEXL22 was expressed and purified as previously reported method [ 17 ]. The HcEXL gene (GenBank accession number ABC30409.1 ) was synthesised by Comate Bioscience Co., Ltd. (Changchun, China).…”

“…Chitin was pretreated with 50 mM sodium acetate buffer (pH 6.0), 5 mg of substrate, and 10 μg of CxEXL22 or HcEXL for 14 h at 50 °C with agitation at 160 rpm. The pH and temperature of the expansin-like proteins were selected based on previous studies [ 17 , 18 ]. The pretreated chitin was washed thrice with distilled water.…”

“…The expansin-like proteins CxEXL22 from Arthrobotrys sp. CX1 and HcEXL from Hahella chejuensis loosen the hydrogen bonds between cellulose polysaccharides, which promotes the accessibility of cellulose and thus enhances cellulose degradation [ 16 , 17 ]. However, there have been few studies on the use of expansin-like proteins to pretreat chitin.…”