万座嗜酸两面菌硫代谢相关膜蛋白基因的筛选

活性巯基在浸矿微生物硫代谢的过程中起着重要的作用,半胱氨酸残基作为蛋白质中活性巯基的提供者,为筛选硫代谢相关蛋白质基因提供了依据。本研究以极端嗜酸热古菌万座嗜酸两面菌Acidianus manzaensis为研究对象,基于其全基因组注释信息,筛选出编码富半胱氨酸残基的潜在硫代谢相关膜蛋白基因,并通过RT-qPCR实验对筛选出来的基因进行表达水平验证,同时利用生物信息学方法对其进一步分析。研究表明,与在亚铁中生长的细胞相比,单质硫培养下的细菌中与能量代谢相关的β-葡糖苷酶,与电子传递相关的ATP合成酶、NADH-辅酶Q氧化还原酶基因均表达上调,说明硫代谢途径可能与能量代谢和电子传递有着重要的联系。此外,还有三个假定蛋白基因表达上调,这三个假定膜蛋白中,ARM75161.1、ARM75436.1中的半胱氨酸都位于保守区域,且均有一个半胱氨酸残基暴露于膜外,而ARM75580.1中的半光氨酸不位于保守区域。其中ARM75436.1具有CXXXC结构域,且该结构域中半胱氨酸残基处于同一个β-折叠中。这些假定蛋白可能参与A. manzaensis中硫代谢途径。

Screening of the Sulfur Metabolism Related Membrane Protein Genes of Acidianus manzaensis

Active thiol groups play an important role in the process of sulfur metabolism of bioleaching microorganisms. As the the provider of active thiol groups in proteins, cysteine residues provide the basis for the screening of proteins, which connected with sulfur metabolism. In this study, the extremely thermophilic archaea Acidianus manzaensis was used as the research object. The potential membrane protein genes related to sulfur metabolism encoding cysteine residues were screened out based on the annotated whole-genome information of A.manzaensis. RT-qPCR was used to verify the expression levels of the screened genes then followed by bioinformatics analysis to further analyze. The results showed that compared with cells grown in ferrous iron, the gene expression of beta-glucosidase related to energy metabolism, ATP synthase and NADH-coenzyme Q oxidoreductase related to the electron transfer in bacteria cultured in elemental sulfur were up-regulated. The results suggested that the sulfur metabolic pathways might be associated with energy metabolism and electron transport. In addition, the expression of three hypothetical protein genes were up-regulated. Among the three hypothetical membrane proteins, the cysteine residues of ARM75161.1 and ARM75436.1 were situated in the conserved domains, and one of the cysteine residues was exposed outside the membrane, while a nother hypothetical protein ARM75580.1 was not located in the conserved region. Among them, the ARM75436.1 had one CXXXC domain, and the cysteine residues in this domain were located in the same β-folding. These putative proteins might participate in the sulfur metabolism pathway in A. manzaensis.