Heterologous expression and periplasmic secretion of an antifungal Bacillus amyloliquefaciensBLB369 endo‐β‐1,3‐1,4‐glucanase in Escherichia coli

Abstract

The endo-β-1,3-1,4-glucanases are glycoside hydrolases involved in the enzymatic depolymerization of 1,3-1,4 β-glucans and showed an antifungal activity against some fungi. Bacillus amyloliquefaciens BLB369 has a high antagonistic activity against phytopathogenic fungi. Its glu369 full-coding sequence of the endo-β-1,3-1,4-glucanase gene (732 bp) was sequenced, cloned and successfully expressed in Escherichia coli Top10. The encoded protein (243 amino acids) has a calculated molecular mass of 27.3 kDa. To simplify the purification procedure, the glu369 coding sequence was cloned into the vector pKJD4. The produced OmpA-His-Glu369 harboured OmpA signal sequence for E. coli periplasmic localization and followed by a 6His residues for its purification. The purified His-tagged proteins revealed two bands on SDS-PAGE analysis with molecular masses of about 30.5 (His-Glu369) and 32.5 kDa (OmpA-His-Glu369).They had the ability to inhibit the growth of phytopathogenic fungus Alternaria alternata. These favourable properties make the endo-β-1,3-1,4-glucanase a good candidate for biotechnological applications. K E Y W O R D Santifungal activity, Bacillus amyloliquefaciens BLB369, endo-β-1,3-1,4-glucanase, heterologous expression

Purification, characterization, immobilization and applications of an enzybiotic β-1,3–1,4-glucanase produced from halotolerant marine Halomonas meridiana ES021

“…A study by Jin et al ( 2011 ) showed that β-1,3–1,4-glucanase from endophytic Bacillus subtilis could be a desirable vital agent against microbial pathogens with higher efficiency and lower toxicity, they proved that the bacterial enzyme had a broad antimicrobial spectrum against fungal and bacterial strains. Antifungal effect of β-1,3–1,4-glucanase was also reported by Dewi et al ( 2016 ), Xu et al ( 2016 ), Zalila-Kolsi et al ( 2018 ) and Yuan et al ( 2020 ).…”

“…(human pathogenic), Yuan et al ( 2020 ) also reported the antifungal effect of β-1,3–1,4-glucanase against Canidia albicans (human pathogenic yeast). The enzyme also showed a weak antifungal effect against the phytopathogenic Aspergillus niger , the antifungal activity of the enzyme against phytopathogenic fungi was previously reported against Curvularia affinis and Colletotrichum gloeosporioides (Dewi et al 2016 ), Cryphonectria parasitica , Cylindrocladium quinqueseptatumI, Helicobasidium purpureum (Xu et al 2016 ), and Alternaria alternata (Zalila‐Kolsi et al 2018 ). Juice clarification by semi-purified β-1,3–1,4-glucanase enzyme showed variation in clarification yields of different juices at their different pH values.…”

“…β-1,3–1,4-glucanase (Ec.3.2.1.73, lichenase) is an industrially important enzyme because of its strict specificity for β-glucan cleavage (Jang et al 2021 ). β-1,3–1,4-glucanase is a glycoside hydrolase involved in β-1,3–1,4-glucans enzymatic depolymerization (Zalila-Kolsi et al 2018 ). It exhibits a strict substrate specificity for cleavage of β-1,4 glycosidic bonds (Sun et al 2012 ).…”

Bacillus amyloliquefaciens: Harnessing Its Potential for Industrial, Medical, and Agricultural Applications—A Comprehensive Review

“…The resultant OmpA-His-Glu369 fusion protein incorporated the OmpA signal sequence for E. coli periplasmic targeting, followed by a 6xHistidine tag for purification purposes. Owing to these beneficial attributes, endo-β-1,3-1,4-glucanase holds significant potential for various biotechnological applications [ 35 ] ( Figure 3 ).…”

“…Furthermore, B. amyloliquefaciens has applications in biotechnology, where it is employed as a host to produce heterologous proteins and as a source of valuable biomolecules. The diverse applications of B. amyloliquefaciens highlight its importance in advancing sustainable and eco-friendly solutions across multiple sectors [ 5 , 8 , 11 , 17 , 21 , 24 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 ].…”