PXD048982-1

PXD048982 is an original dataset announced via ProteomeXchange.

Title	Assessment of biarsenical dye-reactive cysteine residues of human metallothionein MT1E
Description	Metallothioneis are low-molecular weight proteins engaged in zinc and copper ion homeostasis and redox potential regulation of the cell. Their clinical importance lies in their role in cancer resistance to cisplatin related drugs, which is related to their capability of binding sole toxic metal ions, as well as their complexes. Due to metallothioneins' intrinsic properties a satisfactory analytical method to detect, measure and quantify this family of proteins is lacking. In the study project at hand a fluorescent biarsenical probe, CrAsH-EDT2, is presented as specific means for the detection of human metallothioneins. Biarsencial dyes comprise of two arsenic atoms incorporated covalently into a fluorophore structure. Such chemical species show exemplary affinity for thiol groups of cysteine side chains separated in a specific spatial manner; a -CCXXCC- motif being the model one where each arsenic atom is coordinated by two adjacent cysteine thiols. Mass spectrometry analysis was employed in the project in order to pin-point the MT1E metallothionein's cysteine residues responsible for interaction with the CrAsH dye upon pure protein in vitro labelling. Using the extremely tight binding of CrAsH to MT1E to the advantage, a simple approach was undertaken, where the protein was first incubated with the dye at 1:1, 1:2 and 1:3 protein:dye molar ratio, and an alkylation (iodoacetamide (IAA)) and reduction (β-mercaptoethanol (βME)) procedure was carried out on the sample. As controls, dye-untreated non-alkylated and alkylated samples were also processed. Carbamidomethylation of cysteines within MT1E's tryptic peptides was then monitored and used as negative evidence of given cysteine’s participation in dye coordination, assuming the residue as free for the IAA reaction, and thus not occupied by the dye. Previous experiments in the project suggested three possible binding sites of CrAsH within MT1E, hence the analysis comprised of the mentioned three molar ratio steps.
HostingRepository	PRIDE
AnnounceDate	2024-12-16
AnnouncementXML	Submission_2024-12-16_03:50:53.149.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD048982
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Michał Tracz
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	iodoacetamide derivatized residue
Instrument	Synapt MS

Revision	Datetime	Status	ChangeLog Entry
0	2024-01-29 10:23:09	ID requested
⏵ 1	2024-12-16 03:50:53	announced

10.1016/J.ACA.2024.343550;

10.6019/PXD048982;

submitter keyword: alkylation, MSE protein in vitro modification analysis, biarsenical dye, Human, metallothionein, CrAsH,LC-MSE

Michal Tracz
contact affiliation	Laboratory of Protein Mass Spectrometry, Faculty of Biotechnology, University of Wroclaw, Poland
contact email	michal.tracz2@uwr.edu.pl
lab head
Michał Tracz
contact affiliation	Laboratory of Protein Mass Spectrometry, Faculty of Biotechnology, University of Wroclaw
contact email	michal.tracz2@uwr.edu.pl
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD048982
     1. Label: PRIDE project
     2. Name: Assessment of biarsenical dye-reactive cysteine residues of human metallothionein MT1E