Vitor Alison Borges, Farias Keilane Silva, Ribeiro Geise Camila Araújo, Pirovani Carlos Priminho, Benevides Raquel Guimarães, Pereira Gonçalo Amarante Guimarães, de Assis Sandra Aparecida
LAPEM, Biology Department, State University of Feira de Santana, Feira de Santana City, Bahia State Brazil.
Biological Sciences Department, State University of Santa Cruz, Rodovia Jorge Amado, km 16, Ilhéus City, BA 45662-900 Brazil.
3 Biotech. 2024 Nov;14(11):287. doi: 10.1007/s13205-024-04128-x. Epub 2024 Nov 1.
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 expressed in 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 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.
The online version contains supplementary material available at 10.1007/s13205-024-04128-x.
β-葡萄糖苷酶（BGLs）与内切葡聚糖酶和外切葡聚糖酶协同作用，因此在生物质转化为生物乙醇方面具有重要意义。因此，本研究的目的是在细胞中表达产生一种重组β-糖苷酶。在使用麦麸（WB）和羧甲基纤维素（CMC）作为真菌生长培养基后，通过桑格测序确认了酶编码序列的表达。将具有优化密码子用于表达的合成基因betaglyc-GH1克隆到pET-28a中。使用镍柱纯化β-葡萄糖苷酶重组体（GH1嵌合体），并通过质谱确认其身份。重组酶在SDS-PAGE上的表观分子量为53.23 kDa。重组β-葡萄糖苷酶以对硝基苯基-β-D-吡喃葡萄糖苷（pNPG）为底物表现出水解活性，在pH 4.6和65°C时活性最高。因此，结果表明GH1嵌合体在木质纤维素材料水解以获得葡萄糖单体方面的应用可能是有效的。
在线版本包含可在10.1007/s13205-024-04128-x获取的补充材料。
