PXD031169-1

PXD031169 is an original dataset announced via ProteomeXchange.

Title	Engineering and exploiting synthetic allostery of NanoLuc luciferase
Description	The ability to endow constitutively active proteins with synthetic allostery is a central objective of Synthetic Biology, mostly focused on the creation of protein biosensors – artificial sensing proteins with easily quantifiable outputs. Here we hypothesize that circular permutation of proteins increases the probability of functional coupling of new N- and C- terminal sequences with the active center of the protein through increased local structural disorder. We test this by constructing a synthetically allosteric version of circular permutated NanoLuc luciferase, activated through ligand-induced intramolecular non-covalent cyclisation. The developed biosensors cover a range of emission wavelengths and display sensitivity as low as 50 pM and dynamic range as high as 16-fold and could quantify their cognate ligand in human fluids such as saliva, serum and lysed blood. We apply hydrogen exchange kinetic mass spectrometry to analyze time resolved structural changes in the developed biosensors and observed ligand-mediated folding of newly created termini. While a large study is required for determining how frequent synthetic allostery emerges following circular permutation and what types of protein folds are susceptible to it, this study provides motivation and justification for such efforts.
HostingRepository	PRIDE
AnnounceDate	2022-02-10
AnnouncementXML	Submission_2022-02-10_00:57:45.287.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Jonathan Phillips
SpeciesList	scientific name: Escherichia coli; NCBI TaxID: 562;
ModificationList	No PTMs are included in the dataset
Instrument	Synapt MS

Revision	Datetime	Status	ChangeLog Entry
0	2022-01-21 13:23:35	ID requested
⏵ 1	2022-02-10 00:57:45	announced
2	2024-10-22 05:31:11	announced	2024-10-22: Updated project metadata.

Dataset with its publication pending

submitter keyword: LC-MS, HDX, HDMSE, Biosenso, allostery

Jonathan Phillips
contact affiliation	Living Systems Institute, University of Exeter, EX4 4QD, UK
contact email	jj.phillips@exeter.ac.uk
lab head
Jonathan Phillips
contact affiliation	Living Systems Institute, University of Exeter
contact email	jj.phillips@exeter.ac.uk
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/2022/02/PXD031169

PRIDE project URI

• PRIDE
  1. PXD031169
     1. Label: PRIDE project
     2. Name: Engineering and exploiting synthetic allostery of NanoLuc luciferase