{"title":"从内生链霉菌(Streptomyces mobaraensis DB13)中纯化和表征与木薯(Costus speciosus)相关的中等恒温生淀粉消化α-淀粉酶。","authors":"Dina Barman, Mamtaj S Dkhar","doi":"10.2323/jgam.2023.08.001","DOIUrl":null,"url":null,"abstract":"<p><p>Endophytic actinobacteria are known to produce various enzymes with potential industrial applications. Alpha-amylase is an important class of industrial enzyme with a multi-dimensional utility. The present experiment was designed to characterize a moderately thermostable α-amylase producing endophytic Streptomyces mobaraensis DB13 isolated from Costus speciosus (J. Koenig) Sm. The enzyme was purified using 60% ammonium sulphate precipitation, dialysis, and Sephadex G-100 column chromatography. Based on 12% SDS-PAGE, the molecular weight of the purified α-amylase was estimated to be 55 kDa. The maximum α-amylase activity was achieved at pH 7.0, 50°C and it retained 80% of its activity at both pH 7.0 and 8.0 after incubation for 2 h. The α-mylase activity is strongly enhanced by Ca<sup>2+</sup>, Mg<sup>2+</sup>, and inhibited by Ba<sup>2+</sup>. The activity remains stable in the presence of Tween-80, SDS, PMSF, and Triton X-100; however, β-mercaptoethanol, EDTA, and H<sub>2</sub>O<sub>2</sub> reduced the activity. The kinetic parameters K<sub>m</sub> and V<sub>max</sub> values for this α-amylase were calculated as 2.53 mM and 29.42 U/mL respectively. The α-amylase had the ability to digest various raw starches at a concentration of 10 mg/mL at pH 7.0, 50°C, where maize and rice are the preferred substrates. The digestion starts after 4 h of incubation, which reaches maximum after 48 h of incubation. These results suggest that S. mobaraensis DB13 is a potential source of moderately thermostable α-amylase enzyme, that effciently hydrolyzes raw starch. It suggesting that this α-amylase is a promising candidate to be use for industrial purposes.</p>","PeriodicalId":15842,"journal":{"name":"Journal of General and Applied Microbiology","volume":" ","pages":"293-300"},"PeriodicalIF":0.8000,"publicationDate":"2024-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Purification and characterization of moderately thermostable raw-starch digesting α-amylase from endophytic Streptomyces mobaraensis DB13 associated with Costus speciosus.\",\"authors\":\"Dina Barman, Mamtaj S Dkhar\",\"doi\":\"10.2323/jgam.2023.08.001\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Endophytic actinobacteria are known to produce various enzymes with potential industrial applications. Alpha-amylase is an important class of industrial enzyme with a multi-dimensional utility. The present experiment was designed to characterize a moderately thermostable α-amylase producing endophytic Streptomyces mobaraensis DB13 isolated from Costus speciosus (J. Koenig) Sm. The enzyme was purified using 60% ammonium sulphate precipitation, dialysis, and Sephadex G-100 column chromatography. Based on 12% SDS-PAGE, the molecular weight of the purified α-amylase was estimated to be 55 kDa. The maximum α-amylase activity was achieved at pH 7.0, 50°C and it retained 80% of its activity at both pH 7.0 and 8.0 after incubation for 2 h. The α-mylase activity is strongly enhanced by Ca<sup>2+</sup>, Mg<sup>2+</sup>, and inhibited by Ba<sup>2+</sup>. The activity remains stable in the presence of Tween-80, SDS, PMSF, and Triton X-100; however, β-mercaptoethanol, EDTA, and H<sub>2</sub>O<sub>2</sub> reduced the activity. The kinetic parameters K<sub>m</sub> and V<sub>max</sub> values for this α-amylase were calculated as 2.53 mM and 29.42 U/mL respectively. The α-amylase had the ability to digest various raw starches at a concentration of 10 mg/mL at pH 7.0, 50°C, where maize and rice are the preferred substrates. The digestion starts after 4 h of incubation, which reaches maximum after 48 h of incubation. These results suggest that S. mobaraensis DB13 is a potential source of moderately thermostable α-amylase enzyme, that effciently hydrolyzes raw starch. It suggesting that this α-amylase is a promising candidate to be use for industrial purposes.</p>\",\"PeriodicalId\":15842,\"journal\":{\"name\":\"Journal of General and Applied Microbiology\",\"volume\":\" \",\"pages\":\"293-300\"},\"PeriodicalIF\":0.8000,\"publicationDate\":\"2024-05-02\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Journal of General and Applied Microbiology\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.2323/jgam.2023.08.001\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2023/8/25 0:00:00\",\"PubModel\":\"Epub\",\"JCR\":\"Q4\",\"JCRName\":\"BIOTECHNOLOGY & APPLIED MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of General and Applied Microbiology","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.2323/jgam.2023.08.001","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2023/8/25 0:00:00","PubModel":"Epub","JCR":"Q4","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
Purification and characterization of moderately thermostable raw-starch digesting α-amylase from endophytic Streptomyces mobaraensis DB13 associated with Costus speciosus.
Endophytic actinobacteria are known to produce various enzymes with potential industrial applications. Alpha-amylase is an important class of industrial enzyme with a multi-dimensional utility. The present experiment was designed to characterize a moderately thermostable α-amylase producing endophytic Streptomyces mobaraensis DB13 isolated from Costus speciosus (J. Koenig) Sm. The enzyme was purified using 60% ammonium sulphate precipitation, dialysis, and Sephadex G-100 column chromatography. Based on 12% SDS-PAGE, the molecular weight of the purified α-amylase was estimated to be 55 kDa. The maximum α-amylase activity was achieved at pH 7.0, 50°C and it retained 80% of its activity at both pH 7.0 and 8.0 after incubation for 2 h. The α-mylase activity is strongly enhanced by Ca2+, Mg2+, and inhibited by Ba2+. The activity remains stable in the presence of Tween-80, SDS, PMSF, and Triton X-100; however, β-mercaptoethanol, EDTA, and H2O2 reduced the activity. The kinetic parameters Km and Vmax values for this α-amylase were calculated as 2.53 mM and 29.42 U/mL respectively. The α-amylase had the ability to digest various raw starches at a concentration of 10 mg/mL at pH 7.0, 50°C, where maize and rice are the preferred substrates. The digestion starts after 4 h of incubation, which reaches maximum after 48 h of incubation. These results suggest that S. mobaraensis DB13 is a potential source of moderately thermostable α-amylase enzyme, that effciently hydrolyzes raw starch. It suggesting that this α-amylase is a promising candidate to be use for industrial purposes.
期刊介绍:
JGAM is going to publish scientific reports containing novel and significant microbiological findings, which are mainly devoted to the following categories: Antibiotics and Secondary Metabolites; Biotechnology and Metabolic Engineering; Developmental Microbiology; Environmental Microbiology and Bioremediation; Enzymology; Eukaryotic Microbiology; Evolution and Phylogenetics; Genome Integrity and Plasticity; Microalgae and Photosynthesis; Microbiology for Food; Molecular Genetics; Physiology and Cell Surface; Synthetic and Systems Microbiology.