Development of bi-functional chimeric enzyme (CtGH1-L1-CtGH5-F194A) from endoglucanase (CtGH5) mutant F194A and β-1,4-glucosidase (CtGH1) from Clostridium thermocellum with enhanced activity and structural integrity.
Bioethanol
Chimera
Endoglucanase
Site-directed mutagenesis
β-Glucosidase
Journal
Bioresource technology
ISSN: 1873-2976
Titre abrégé: Bioresour Technol
Pays: England
ID NLM: 9889523
Informations de publication
Date de publication:
Jun 2019
Jun 2019
Historique:
received:
11
01
2019
revised:
08
03
2019
accepted:
09
03
2019
pubmed:
22
3
2019
medline:
17
9
2019
entrez:
22
3
2019
Statut:
ppublish
Résumé
Site-directed mutagenesis of β-1,4-endoglucanase from family 5 glycoside hydrolase (CtGH5) from Clostridium thermocellum was performed to develop a mutant CtGH5-F194A that gave 40 U/mg specific activity against carboxymethyl cellulose, resulting 2-fold higher activity than wild-type CtGH5. CtGH5-F194A was fused with a β-1,4-glucosidase, CtGH1 from Clostridium thermocellum to develop a chimeric enzyme. The chimera (CtGH1-L1-CtGH5-F194A) expressed as a soluble protein using E. coli BL-21cells displaying 3- to 5-fold higher catalytic efficiency for endoglucanase and β-glucosidase activities. TLC analysis of hydrolysed product of CMC by chimera 1 revealed glucose as final product confirming both β-1,4-endoglucanase and β-1,4-glucosidase activities, while the products of CtGH5-F194A were cellobiose and cello-oligosaccharides. Protein melting studies of CtGH5-F194A showed melting temperature (T
Identifiants
pubmed: 30897487
pii: S0960-8524(19)30400-6
doi: 10.1016/j.biortech.2019.03.051
pii:
doi:
Substances chimiques
Recombinant Fusion Proteins
0
Cellobiose
16462-44-5
beta-Glucosidase
EC 3.2.1.21
Cellulase
EC 3.2.1.4
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
494-501Informations de copyright
Copyright © 2019 Elsevier Ltd. All rights reserved.