PXD045793

PXD045793 is an original dataset announced via ProteomeXchange.

Title	Post-translationally modified and stop codon readthrough of recombinant Francisella tularensis Universal stress protein in Escherichia coli
Description	Recombinant Francisella tularensis universal stress protein with a C-terminal histidine-tag (rUsp/His6) was expressed in Escherichia coli. Endogenous F. tularensis Usp has a predicted molecular mass of 30 kDa, but rUsp/His6 had an apparent molecular weight of 33 kDa based on Western blot analyses. To determine the source of the higher molecular weight for rUsp/His6, post translational modifications were examined. Tryptic peptides of purified rUsp/His6 were subjected to liquid chromatography tandem mass spectrometry (LC-MS/MS) and fragmentation spectra were searched for acetylated lysines and polyaminated glutamines. Of the 24 lysines in rUsp/His6, 10 were acetylated (K63, K68, K72, K129, K175, K201, K208, K212, K233, and K238) and three of the four glutamines had putrescine, spermidine and spermine adducts (Q55, Q60 and Q267). The level of post-translational modification was substoichiometric, eliminating the possibility that these modifications were the sole contributor to the 3 kDa extra mass of rUsp/His6. LC-MS/MS revealed that stop codon readthrough had occurred resulting in the unexpected addition of 20 extra amino acids at the C-terminus of rUsp/His6, after the histidine tag. Further, the finding of polyaminated glutamines in rUsp/His6 indicated that E. coli is capable of transglutaminase activity.
HostingRepository	PRIDE
AnnounceDate	2024-05-24
AnnouncementXML	Submission_2024-05-24_01:08:18.276.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Lawrence Schopfer
SpeciesList	scientific name: Francisella tularensis subsp. tularensis; NCBI TaxID: 119856;
ModificationList	acetylated residue; iodoacetamide derivatized residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2023-09-28 15:13:28	ID requested
⏵ 1	2024-05-24 01:08:19	announced

10.1371/journal.pone.0299701;

Girardo B, Schopfer LM, Yue Y, Lockridge O, Larson MA, Polyaminated, acetylated and stop codon readthrough of recombinant Francisella tularensis universal stress protein in Escherichia coli. PLoS One, 19(4):e0299701(2024) [pubmed]

submitter keyword: Escherichia coli pET expression system, acetylation, Francisella tularensis Universal stress protein, transglutaminase, polyamination, stop codon readthrough

Marilynn A. Larson
contact affiliation	Pathology and Microbiology Department, University of Nebraska Medical Center, Omaha, NE, USA
contact email	malarson@unmc.edu
lab head
Lawrence Schopfer
contact affiliation	University of Nebraska Medical Center
contact email	lmschopf@unmc.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD045793
     1. Label: PRIDE project
     2. Name: Post-translationally modified and stop codon readthrough of recombinant Francisella tularensis Universal stress protein in Escherichia coli