Expression and functional characterization of a low-temperature glucanase derived from rumen microbiota of Hu sheep
Glucanase is an important additive with significant application value in feed,food,and textile industries.In this study,the IDSGH5-50 gene was amplified from the rumen microbiota cDNA of Hu sheep and heterologously expressed in Escherichia coli BL21(DE3)to investigate its enzymatic properties and hydrolysis products.The results revealed that IDSGH5-50 encoded 690 amino acids,with a molecular weight of 75.78 kDa and an isoelectric point of 4.75.The optimal reaction conditions for rIDSGH5-50 were 30℃and pH 6.0,and it maintained high activity(＞70%)at temperatures of 4-20℃.It exhibited stable activity at pH 5.0-8.0.The active substrate spectrum of rIDSGH5-50 showed that rIDSGH5-50 could catalyze barley β-glucan,lichenin,tamarind xyloglucan,and konjac gum,and the specific activities of them were(10.42±0.16),(7.12±0.08),(7.03±0.38),and(5.94±0.65)U/mg,respectively.Thin layer chromatography analysis showed that barley β-glucan was mainly degraded to cellotetrose and cellopentose,that lichenin was mainly degraded to cellotriose and cellopentose,that konjac gum was mainly degraded to cellopentose,and that tamarind xyloglucan was mainly degraded to oligosaccharides with a degree of polymerization＞5,which was catalyzed by rIDSGH5-50.This study reported a cold-adapted β-1,4-glucanase IDSGH5-50 from the genus Ruminococcus,establishing the foundation for enzyme preparations used in feed and food enzyme preparation.
Hu sheep rumenglucanaselow temperaturecold adaptationsubstrate hydrolysis
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