Table_1_Proteomics unveil a central role for peroxisomes in butyrate assimilation of the heterotrophic Chlorophyte alga Polytomella sp..XLSX

Volatile fatty acids found in effluents of the dark fermentation of biowastes can be used for mixotrophic growth of microalgae, improving productivity and reducing the cost of the feedstock. Microalgae can use the acetate in the effluents very well, but butyrate is poorly assimilated and can inhibit growth above 1 gC.L−1. The non-photosynthetic chlorophyte alga Polytomella sp. SAG 198.80 was found to be able to assimilate butyrate fast. To decipher the metabolic pathways implicated in butyrate assimilation, quantitative proteomics study was developed comparing Polytomella sp. cells grown on acetate and butyrate at 1 gC.L−1. After statistical analysis, a total of 1772 proteins were retained, of which 119 proteins were found to be overaccumulated on butyrate vs. only 46 on acetate, indicating that butyrate assimilation necessitates additional metabolic steps. The data show that butyrate assimilation occurs in the peroxisome via the β-oxidation pathway to produce acetyl-CoA and further tri/dicarboxylic acids in the glyoxylate cycle. Concomitantly, reactive oxygen species defense enzymes as well as the branched amino acid degradation pathway were strongly induced. Although no clear dedicated butyrate transport mechanism could be inferred, several membrane transporters induced on butyrate are identified as potential condidates. Metabolic responses correspond globally to the increased needs for central cofactors NAD, ATP and CoA, especially in the peroxisome and the cytosol.

生物废弃物暗发酵废液中所含的挥发性脂肪酸（Volatile fatty acids）可用于微藻（microalgae）的混养生长，既能提升培养产率，又可降低原料成本。微藻对废液中的乙酸盐（acetate）利用效果良好，但丁酸盐（butyrate）的同化能力较弱，且当浓度超过1 gC·L⁻¹时会抑制微藻生长。研究发现非光合绿藻（non-photosynthetic chlorophyte alga）*Polytomella* sp. SAG 198.80可快速同化丁酸盐。为解析与丁酸盐同化相关的代谢通路，本研究开展了定量蛋白质组学（quantitative proteomics）研究，对比了分别以1 gC·L⁻¹浓度的乙酸盐和丁酸盐为碳源培养的*Polytomella* sp.细胞。经统计分析，最终共鉴定到1772种蛋白质；其中，在丁酸盐培养条件下上调富集的蛋白质共119种，而乙酸盐组仅为46种，这表明丁酸盐同化需要额外的代谢步骤。实验数据表明，丁酸盐同化过程发生于过氧化物酶体（peroxisome）中，通过β-氧化途径（β-oxidation pathway）生成乙酰辅酶A（acetyl-CoA），随后经乙醛酸循环（glyoxylate cycle）生成三羧酸/二羧酸类物质。与此同时，活性氧（reactive oxygen species）防御酶类以及支链氨基酸降解通路均被显著诱导表达。尽管目前无法明确推断出专一性的丁酸盐转运机制，但本研究鉴定出若干在丁酸盐培养条件下上调的膜转运蛋白，可作为潜在的候选转运蛋白。整体而言，该代谢响应与核心辅因子烟酰胺腺嘌呤二核苷酸（NAD）、三磷酸腺苷（ATP）及辅酶A（CoA）的需求提升相契合，这一现象在过氧化物酶体与细胞质（cytosol）中尤为明显。