RAW Data PhyscoGlcStlproteome

Differential protein expression in response to glucose in the moss Physcomitrella patens was evaluated, for this 10-day old protonemata were exposed to 0 and 300 mM of glucose for 24 h. The protonemata were frozen in liquid nitrogen and the total proteins were extracted and the concentration was determined with the Bradford method and the protein quality and quantity were verified by SDS-PAGE. Total proteins from three biological replicates were reduced with dithiothreitol (DTT), alkylated with iodoacetamide (Sigma-Aldrich), and digested with trypsin ((Promega Modified Trypsin Sequencing Grade). The resulting peptides were applied to a pump LC-MS nanoflow EASY-nLC II instrument coupled to a mass spectrometer LTQ Orbitrap-Velos system with nano-electrospray ionization (Thermo-Fisher Scientific Co.,San Jose, CA). All MS/MS samples from three biological replicates were analyzed using Sequest and X! Tandem for peptide identification. Both tools were set up to search on the uniprot-physcomitrella+patens.fasta file (UP000006727, 35539 entries) assuming trypsin digestion. Sequest and X! Tandem were used considering a fragment ion mass tolerance of 0.60 Da and a parent ion tolerance of 20 ppm. Cysteine carbamidomethyl was considered as a fixed modification, whereas histidine carbamidomethyl, methionine oxidation, and Glu- >pyro-Glu, Gln->pyro-Glu and ammonia-loss of the n-terminus were specified as variable modifications. Protein identification from the three biological replicates was carried out using the software tool Scaffold. These procedures were carried out at the Proteomic unit from Universidad Nacional Autónoma de México.

在苔藓植物 Physcomitrella patens 中，对葡萄糖刺激下的差异蛋白质表达进行了评估。对于10天的原丝体，将其暴露于0和300 mM的葡萄糖溶液中，持续24小时。原丝体被液氮冷冻，总蛋白质被提取，并使用Bradford方法测定其浓度。通过SDS-PAGE验证了蛋白质的质量和数量。来自三个生物学重复的总蛋白质样品使用二硫苏糖醇（DTT）进行还原，用碘代乙酰胺（Sigma-Aldrich）进行烷基化，并使用蛋白酶（Promega Modified Trypsin Sequencing Grade）进行消化。所得肽段被应用于连接质谱仪LTQ Orbitrap-Velos系统的LC-MS纳米流EASY-nLC II泵。所有来自三个生物学重复的MS/MS样品均使用Sequest和X! Tandem进行肽段鉴定。这两个工具被设置为在uniprot-physcomitrella+patens.fasta文件（UP000006727，35539条条目）上进行搜索，假设进行了蛋白酶消化。考虑到片段离子质量容差为0.60 Da，母离子容差为20 ppm。半胱氨酸碳酰胺甲基化被作为固定修饰，而组氨酸碳酰胺甲基化、蛋氨酸氧化以及谷氨酰>吡咯谷氨酸、谷氨酰>吡咯谷氨酸和N端氨损失被指定为可变修饰。使用Scaffold软件工具对三个生物学重复的蛋白质进行鉴定。上述过程在墨西哥国立自治大学的蛋白质组学单元进行。