{"title":"源自 Moniliophthora perniciosa (Stahel) Aime 和 Phillips Mora 的 β-葡萄糖苷酶的克隆、异源表达和特征描述。","authors":"Alison Borges Vitor, Keilane Silva Farias, Geise Camila Araújo Ribeiro, Carlos Priminho Pirovani, Raquel Guimarães Benevides, Gonçalo Amarante Guimarães Pereira, Sandra Aparecida de Assis","doi":"10.1007/s13205-024-04128-x","DOIUrl":null,"url":null,"abstract":"<p><p>Β-glucosidase (BGLs) act synergistically with endoglucanases and exoglucanases and then are of great interest for biomass conversion into bioethanol. Thus, the aim of the current study is to produce a recombinant β-glycosidase from <i>Moniliophtora perniciosa</i> expressed in <i>Escherichia coli</i> cells. Enzyme coding sequence expression was confirmed through Sanger sequencing after using wheat bran (WB) and carboxymethylcellulose (CMC) as fungal growth media. Synthetic gene betaglyc-GH1 with optimized codons for <i>E. coli</i> expression was cloned in pET-28a. β-glucosidase recombinant (GH1chimera) was purified using a nickel column and its identity was confirmed through mass spectrometry. The recombinant enzyme presented an apparent molecular mass of 53.23 kDa on SDS-PAGE. Recombinant β-glucosidase has shown hydrolytic activity using p-nitrophenyl-β-D-glycopyranoside (pNPG) as substrate and maximum activity at pH 4.6 and 65 °C. Thus, the results indicate that the application of the GH1chimera in the hydrolysis of lignocellulosic materials to obtain glucose monomers can be efficient.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s13205-024-04128-x.</p>","PeriodicalId":7067,"journal":{"name":"3 Biotech","volume":null,"pages":null},"PeriodicalIF":2.6000,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11530418/pdf/","citationCount":"0","resultStr":"{\"title\":\"Cloning, heterologous expression and characterization of β-glucosidase deriving from <i>Moniliophthora perniciosa</i> (Stahel) Aime and Phillips Mora.\",\"authors\":\"Alison Borges Vitor, Keilane Silva Farias, Geise Camila Araújo Ribeiro, Carlos Priminho Pirovani, Raquel Guimarães Benevides, Gonçalo Amarante Guimarães Pereira, Sandra Aparecida de Assis\",\"doi\":\"10.1007/s13205-024-04128-x\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Β-glucosidase (BGLs) act synergistically with endoglucanases and exoglucanases and then are of great interest for biomass conversion into bioethanol. Thus, the aim of the current study is to produce a recombinant β-glycosidase from <i>Moniliophtora perniciosa</i> expressed in <i>Escherichia coli</i> cells. Enzyme coding sequence expression was confirmed through Sanger sequencing after using wheat bran (WB) and carboxymethylcellulose (CMC) as fungal growth media. Synthetic gene betaglyc-GH1 with optimized codons for <i>E. coli</i> expression was cloned in pET-28a. β-glucosidase recombinant (GH1chimera) was purified using a nickel column and its identity was confirmed through mass spectrometry. The recombinant enzyme presented an apparent molecular mass of 53.23 kDa on SDS-PAGE. Recombinant β-glucosidase has shown hydrolytic activity using p-nitrophenyl-β-D-glycopyranoside (pNPG) as substrate and maximum activity at pH 4.6 and 65 °C. Thus, the results indicate that the application of the GH1chimera in the hydrolysis of lignocellulosic materials to obtain glucose monomers can be efficient.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s13205-024-04128-x.</p>\",\"PeriodicalId\":7067,\"journal\":{\"name\":\"3 Biotech\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":2.6000,\"publicationDate\":\"2024-11-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11530418/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"3 Biotech\",\"FirstCategoryId\":\"5\",\"ListUrlMain\":\"https://doi.org/10.1007/s13205-024-04128-x\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"BIOTECHNOLOGY & APPLIED MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"3 Biotech","FirstCategoryId":"5","ListUrlMain":"https://doi.org/10.1007/s13205-024-04128-x","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
Cloning, heterologous expression and characterization of β-glucosidase deriving from Moniliophthora perniciosa (Stahel) Aime and Phillips Mora.
Β-glucosidase (BGLs) act synergistically with endoglucanases and exoglucanases and then are of great interest for biomass conversion into bioethanol. Thus, the aim of the current study is to produce a recombinant β-glycosidase from Moniliophtora perniciosa expressed in Escherichia coli cells. Enzyme coding sequence expression was confirmed through Sanger sequencing after using wheat bran (WB) and carboxymethylcellulose (CMC) as fungal growth media. Synthetic gene betaglyc-GH1 with optimized codons for E. coli expression was cloned in pET-28a. β-glucosidase recombinant (GH1chimera) was purified using a nickel column and its identity was confirmed through mass spectrometry. The recombinant enzyme presented an apparent molecular mass of 53.23 kDa on SDS-PAGE. Recombinant β-glucosidase has shown hydrolytic activity using p-nitrophenyl-β-D-glycopyranoside (pNPG) as substrate and maximum activity at pH 4.6 and 65 °C. Thus, the results indicate that the application of the GH1chimera in the hydrolysis of lignocellulosic materials to obtain glucose monomers can be efficient.
Supplementary information: The online version contains supplementary material available at 10.1007/s13205-024-04128-x.
3 BiotechAgricultural and Biological Sciences-Agricultural and Biological Sciences (miscellaneous)
CiteScore
6.00
自引率
0.00%
发文量
314
期刊介绍:
3 Biotech publishes the results of the latest research related to the study and application of biotechnology to:
- Medicine and Biomedical Sciences
- Agriculture
- The Environment
The focus on these three technology sectors recognizes that complete Biotechnology applications often require a combination of techniques. 3 Biotech not only presents the latest developments in biotechnology but also addresses the problems and benefits of integrating a variety of techniques for a particular application. 3 Biotech will appeal to scientists and engineers in both academia and industry focused on the safe and efficient application of Biotechnology to Medicine, Agriculture and the Environment.