RAW Data PhyscoGlcStlproteome

Published: 16-11-2020| Version 1 | DOI: 10.17632/t5m28m66vc.1
Contributors:
Alejandra Chamorro-Flores,
Miguel Angel Villalobos-López,
Angel Arturo Guevara-García,
Analilia Arroyo-Becerra

Description

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.

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