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从里氏木霉中鉴定出一种具有高度特异性的 L-阿拉伯糖转运蛋白。

Functional characterization of a highly specific L-arabinose transporter from Trichoderma reesei.

机构信息

VTT Technical Research Center of Finland Ltd, Tietotie 2, 02150, Espoo, Finland.

Membrane Transport Discovery Lab, Department of Nephrology and Hypertension, University of Bern, Freiburgstrasse 15, 3010, Bern, Switzerland.

出版信息

Microb Cell Fact. 2021 Sep 8;20(1):177. doi: 10.1186/s12934-021-01666-4.

Abstract

BACKGROUND

Lignocellulose biomass has been investigated as a feedstock for second generation biofuels and other value-added products. Some of the processes for biofuel production utilize cellulases and hemicellulases to convert the lignocellulosic biomass into a range of soluble sugars before fermentation with microorganisms such as yeast Saccharomyces cerevisiae. One of these sugars is L-arabinose, which cannot be utilized naturally by yeast. The first step in L-arabinose catabolism is its transport into the cells, and yeast lacks a specific transporter, which could perform this task.

RESULTS

We identified Trire2_104072 of Trichoderma reesei as a potential L-arabinose transporter based on its expression profile. This transporter was described already in 2007 as D-xylose transporter XLT1. Electrophysiology experiments with Xenopus laevis oocytes and heterologous expression in yeast revealed that Trire2_104072 is a high-affinity L-arabinose symporter with a K value in the range of [Formula: see text] 0.1-0.2 mM. It can also transport D-xylose but with low affinity (K [Formula: see text] 9 mM). In yeast, L-arabinose transport was inhibited slightly by D-xylose but not by D-glucose in an assay with fivefold excess of the inhibiting sugar. Comparison with known L-arabinose transporters revealed that the expression of Trire2_104072 enabled yeast to uptake L-arabinose at the highest rate in conditions with low extracellular L-arabinose concentration. Despite the high specificity of Trire2_104072 for L-arabinose, the growth of its T. reesei deletion mutant was only affected at low L-arabinose concentrations.

CONCLUSIONS

Due to its high affinity for L-arabinose and low inhibition by D-glucose or D-xylose, Trire2_104072 could serve as a good candidate for improving the existing pentose-utilizing yeast strains. The discovery of a highly specific L-arabinose transporter also adds to our knowledge of the primary metabolism of T. reesei. The phenotype of the deletion strain suggests the involvement of other transporters in L-arabinose transport in this species.

摘要

背景

木质纤维素生物质已被研究作为第二代生物燃料和其他增值产品的原料。一些生物燃料生产过程利用纤维素酶和半纤维素酶将木质纤维素生物质转化为一系列可发酵微生物(如酵母酿酒酵母)的可溶性糖。这些糖之一是 L-阿拉伯糖,酵母不能自然利用。L-阿拉伯糖分解代谢的第一步是将其运输到细胞内,而酵母缺乏能够完成此任务的特定转运蛋白。

结果

我们根据其表达谱确定里氏木霉中的 Trire2_104072 为潜在的 L-阿拉伯糖转运蛋白。该转运蛋白早在 2007 年就被描述为 D-木糖转运蛋白 XLT1。用非洲爪蟾卵母细胞进行电生理学实验和在酵母中的异源表达表明,Trire2_104072 是一种高亲和力的 L-阿拉伯糖协同转运蛋白,K 值在 [公式:见正文] 0.1-0.2 mM 范围内。它还可以运输 D-木糖,但亲和力较低(K [公式:见正文] 9 mM)。在酵母中,L-阿拉伯糖的运输在 5 倍过量抑制糖的测定中被 D-木糖轻微抑制,但不受 D-葡萄糖的抑制。与已知的 L-阿拉伯糖转运蛋白的比较表明,在低细胞外 L-阿拉伯糖浓度条件下,Trire2_104072 的表达使酵母能够以最高的速率摄取 L-阿拉伯糖。尽管 Trire2_104072 对 L-阿拉伯糖具有高度特异性,但 T. reesei 缺失突变体的生长仅在低 L-阿拉伯糖浓度下受到影响。

结论

由于其对 L-阿拉伯糖的高亲和力和对 D-葡萄糖或 D-木糖的低抑制作用,Trire2_104072 可以作为改善现有戊糖利用酵母菌株的良好候选物。高度特异性 L-阿拉伯糖转运蛋白的发现也增加了我们对里氏木霉初级代谢的认识。缺失菌株的表型表明,其他转运蛋白也参与了该物种的 L-阿拉伯糖运输。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/3ba9/8425032/fba68ef2800c/12934_2021_1666_Fig1_HTML.jpg

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