PXD015549

PXD015549 is an original dataset announced via ProteomeXchange.

Title	LCMS quantification of internal crosslink in WT and G68L mutant R2lox proteins from Geobacillus kaustophilus.
Description	A heterobimetallic Mn/Fe cofactor is found in the R2 subunit of class Ic ribonucleotide reductases (R2c) and R2-like ligand-binding oxidases (R2lox). Although the protein-derived metal ligands are the same in both groups of proteins, the connectivity of the two metal ions and the chemistry each cofactor performs are different: in R2c, a one-electron oxidant, the Mn/Fe dimer is linked by two oxygen bridges (μ-oxo/μ-hydroxo), whereas in R2lox, a two-electron oxidant, it is linked by a single oxygen bridge (μ-hydroxo) and a fatty acid ligand. Here we identify a second coordination sphere residue which directs the divergent reactivity of the protein scaffold. The residue that directly precedes the N-terminal carboxylate metal ligand is conserved as a glycine within the R2lox group, but not in R2c. Mutation of the glycine to leucine converts the resting state R2lox cofactor into an R2c-like cofactor, a µ-oxo/µ-hydroxo bridged MnIII/FeIII dimer. This species has recently been observed as an intermediate of the oxygen activation reaction in wild-type R2lox, demonstrating that it is physiologically relevant. Cofactor maturation in R2c and R2lox therefore follows the same pathway, with structural and functional divergence of the two cofactor forms following oxygen activation. We also show that the leucine mutant no longer functions as a two-electron oxidant. Specifically the mass spectrometry data here deposited tracks the abundance of the cross-linked peptide AVIRAATVYNMIVE-AVTLD, which was used as surrogate marker for the abundance of the internal V72-Y162 ether cross-link, which is directly impacted by the metalation state of the protein as well as the residue present in position 68 (G or L). Taken together, our data demonstrate that the residue preceding the N-terminal metal ligand directs the cofactor’s reactivity towards one- or two-electron redox chemistry, presumably by setting the protonation state of the bridging oxygens and thereby perturbing the redox potential of the Mn ion.
HostingRepository	PRIDE
AnnounceDate	2019-09-26
AnnouncementXML	Submission_2019-09-26_06:52:09.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Rui Branca
SpeciesList	scientific name: Geobacillus kaustophilus; NCBI TaxID: 1462;
ModificationList	No PTMs are included in the dataset
Instrument	LTQ Orbitrap Velos

Revision	Datetime	Status	ChangeLog Entry
0	2019-09-23 04:02:50	ID requested
1	2019-09-26 06:15:44	announced
⏵ 2	2019-09-26 06:52:10	announced	2019-09-26: Updated publication reference for PubMed record(s): 25153930.

Shafaat HS, Griese JJ, Pantazis DA, Roos K, Andersson CS, Popovi, ć, -Bijeli, ć A, Gr, ä, slund A, Siegbahn PE, Neese F, Lubitz W, H, ö, gbom M, Cox N, Electronic structural flexibility of heterobimetallic Mn/Fe cofactors: R2lox and R2c proteins. J Am Chem Soc, 136(38):13399-409(2014) [pubmed]

submitter keyword: ferritin

metalloprotein

ribonucleotide reductase

R2-like ligand-binding oxidase

R2lox

crosslink

Janne Lehtiö
contact affiliation	Dept. Oncology Pathology, Karolinska Institutet and Scilifelab, Stockholm, Sweden
contact email	janne.lehtio@ki.se
lab head
Rui Branca
contact affiliation	Clinical Proteomics Unit, Dep. of Oncology-Pathology
contact email	rui.mamede-branca@ki.se
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2019/09/PXD015549

PRIDE project URI

• PRIDE
  1. PXD015549
     1. Label: PRIDE project
     2. Name: LCMS quantification of internal crosslink in WT and G68L mutant R2lox proteins from Geobacillus kaustophilus.