A novel versatile glycoside hydrolase Cel5M was identified from Pectobacterium carotovorum and used in ramie degumming. The full-length Cel5M (GenBank: MH544226) containing the catalytic domain Cel5MD and its carbohydrate-binding module (CBM3) were expressed in E. coli. Both Cel5M and Cel5MD showed a broad substrate spectrum cleaving konjac glucomannan (KGM), carboxymethylcellulose (CMC) and oat spelt xylan. The optimal temperature of Cel5MD was 55oC with CMC and KGM, whereas the optimum temperature of Cel5M was 65oC with KGM and 60oC with CMC. The improved thermal activity with KGM was probably reflected also in the much lower Km with KGM than with CMC, indicating stronger binding of the full-length enzyme with KGM. Both enzymes were highly stable over a broad pH range from pH 5.0 to 9.0 with KGM. The use of the Cel5M enzyme together with the wild-type P. carotovorum HG-49 strain showed significant improvement when compared to using the mere strain HG-49 for application in ramie degumming. Moreover, the whole P. carotovorum Cel5M gene was over-expressed in P. carotovorum HG-49 itself to improve the ramie-degumming capacity. Altogether, this study indicates that in situ production of processing enzymes could be utilized in the textile industry.
GardettiMAMuthuSS. Handbook of sustainable luxury textiles and fashion, Singapore: Springer Science+Business Media, 2016.
2.
BiswasDChakrabartiSKDeS, et al.Eco-friendly degumming technology for ramie fiber. J Nat Fibers2016; 13: 227–237.
3.
FanPHeFYangY, et al.In-situ microbial degumming technology with Bacillus sp. HG-28 for industrial production of ramie fibers. Biochem Eng J2015; 97: 50–58.
4.
AspeborgHCoutinhoPMWangY, et al.Evolution, substrate specificity and subfamily classification of glycoside hydrolase family 5 (GH5). BMC Evol Biol2012; 12: 186.
5.
ChenZWFriedlandGDPereiraJH, et al.Tracing determinants of dual substrate specificity in glycoside hydrolase family 5. J Biol Chem2012; 287: 25335–25343.
6.
CantarelBLCoutinhoPMRancurelC, et al.The Carbohydrate-Active EnZymes database (CAZy): an expert resource for glycogenomics. Nucleic Acids Res2009; 37: D233–D238.
7.
ZhengFTuTWangXY, et al.Enhancing the catalytic activity of a novel GH5 cellulase GtCel5 from Gloeophyllum trabeum CBS 900.73 by site-directed mutagenesis on loop 6. Biotechnol Biofuels2018; 11: 76.
8.
McGregorNMorarMFengerTH, et al.Structure-function analysis of a mixed-linkage β-glucanase/xyloglucanase from the key ruminal bacteroidetes Prevotella bryantii B(1)4. J Biol Chem2016; 291: 1175–1197.
Reyes-OrtizVHeinsRAChengG, et al.Addition of a carbohydrate-binding module enhances cellulase penetration into cellulose substrates. Biotechnol Biofuels2013; 6: 93.
11.
OliveiraCSepúlvedaGAguiarTQ, et al.Modification of paper properties using carbohydrate-binding module 3 from the Clostridium thermocellum CipA scaffolding protein produced in Pichia pastoris: elucidation of the glycosylation effect. Cellulose2015; 22: 2755–2765.
12.
DuanSWChengLFLiuZC, et al.Diversity and characteristics of kenaf bast degumming microbial resources. J Nat Fibers2018; 15: 799–807.
13.
MaisuriaVBNerurkarAS. Biochemical properties and thermal behavior of pectate lyase produced by Pectobacterium carotovorum subsp. carotovorum BR1 with industrial potentials. Biochem Eng J2012; 63: 22–30.
14.
CamiloCMPolikarpovI. High-throughput cloning, expression and purification of glycoside hydrolases using Ligation-Independent Cloning (LIC). Protein Expres Purif2014; 99: 35–42.
15.
BiasiniMBienertSWaterhouseA, et al.SWISS-MODEL: modelling protein tertiary and quaternary structure using evolutionary information. Nucleic Acids Res2014; 42: W252–W258.
16.
WangYWShiPJLuoHY, et al.Cloning, over-expression and characterization of an alkali-tolerant endo-β-1,4-mannanase from Penicillium freii F63. J Biosci Bioeng2012; 113: 710–714.
17.
VarrotASchüleinMCottazS, et al.Insights into ligand-induced conformational change in Cel5A from Bacillus agaradhaerens revealed by a catalytically active crystal form. J Mol Biol2000; 297: 819–828.
18.
PetkunSJindouSShimonJW, et al.Structure of a family 3b′ carbohydrate-binding module from the Cel9V glycoside hydrolase from Clostridium thermocellum: structural diversity and implications for carbohydrate binding. Acta crystallogr D2010; 66: 33–43.
19.
FortSVarrotASchüleinM, et al.Mixed-linkage cellooligosaccharides: a new class of glycoside hydrolase inhibitors. Chembiochem2001; 2: 319–325.
20.
AngeliniLGScalabrelliMTavariniS, et al.Ramie fibers in a comparison between chemical and microbiological retting proposed for application in biocomposites. Ind Crop Prod2015; 75: 178–184.
21.
WangYWShuTFanP, et al.Characterization of a recombinant alkaline thermostable β-mannanase and its application in eco-friendly ramie degumming. Process Biochem2017; 61: 73–79.
22.
LiZFLiZLDingRY, et al.Composition of ramie hemicelluloses and effect of polysaccharides on fiber properties. Text Res J2016; 86: 451–460.
23.
ChengYLiuZCZengJ, et al.Construction and co-expression of polycistronic plasmids encoding bio-degumming-related enzymes to improve the degumming process of ramie fibres. Biotechnol Lett2016; 38: 2089–2096.
24.
WeiJSongYLiuSY, et al.A green degumming process of ramie. Ind Crop Prod2018; 120: 131–134.
25.
BeraAGhoshAMukhopadhyayA, et al.Improvement of degummed ramie fiber properties upon treatment with cellulase secreting immobilized A. larrymoorei A1. Bioproc Biosyst Eng2015; 38: 341–351.
26.
XiongHRFenelFLeisolaM, et al.Engineering the thermostability of Trichoderma reesei endo-1,4-β-xylanase II by combination of disulphide bridges. Extremophiles2004; 8: 393–400.
27.
ParkYWLimSTChoSJ, et al.Characterization of Erwinia carotovora subsp. carotovora LY34 endo-1,4-β-glucanase genes and rapid identification of their gene products. Biochem Biophys Res Commun1997; 241: 636–641.
28.
CooperVJCSalmondGPC. Molecular analysis of the major cellulase (CelV) of Erwinia carotovora: evidence for an evolutionary “mix-and-match” of enzyme domains. Mol Gen Genet1993; 241: 341–350.
29.
OlsenOThomsenKKWeberJ, et al.Transplanting two unique beta-glucanase catalytic activities into one multienzyme, which forms glucose. Biotechnology1996; 14: 71–76.
30.
MorrillJKulcinskajaESulewskaAM, et al.The GH5 1,4-β-mannanase from Bifidobacterium animalis subsp. lactis Bl-04 possesses a low-affinity mannan-binding module and highlights the diversity of mannanolytic enzymes. BMC Biochem2015; 16: 26.
31.
HervéCRogowskiABlakeAW, et al.Carbohydrate-binding modules promote the enzymatic deconstruction of intact plant cell walls by targeting and proximity effects. Proc Natl Acad Sci2010; 107: 15293–15298.
32.
Ordaz-OrtizJJMarcusSEPaulKJ. Cell wall microstructure analysis implicates hemicellulose polysaccharides in cell adhesion in tomato fruit pericarp parenchyma. Mol Plant2009; 2: 910–921.
33.
WangQChenHGFangG, et al.Isolation of Bacillus cereus P05 and Pseudomonas sp. X12 and their application in the ramie retting. Ind Crop Prod2017; 97: 518–524.
