NMR paper Multiple Site-Specific Phosphorylation of IDPs Monitored by NMR.

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[NMR paper] Multiple Site-Specific Phosphorylation of IDPs Monitored by NMR.

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NMR processing:

MDD

NMR assignment:

Backbone:

Side-chains:

NOEs:

Structure from NMR restraints:

Ab initio:

Fragment-based:

Rosetta-NMR (Robetta)

Template-based:

GeNMR

I-TASSER

Refinement:

Amber

Structure from chemical shifts:

Fragment-based:

Homology-based:

Torsion angles from chemical shifts:

Preditor

TALOS

Promega- Proline

Secondary structure from chemical shifts:

CSI (via RCI server)

TALOS

MICS caps, β-turns

d2D

PECAN

Flexibility from chemical shifts:

RCI

Interactions from chemical shifts:

HADDOCK

Chemical shifts re-referencing:

NMR model quality:

NOEs, other restraints:

Chemical shifts:

RDCs:

Pseudocontact shifts:

Protein geomtery:

SAVES2 or SAVES4

Quality Control Check

NMR spectrum prediction:

FANDAS

MestReS

V-NMR

Flexibility from structure:

Backbone S2

Methyl S2

B-factor

Molecular dynamics:

Gromacs

Amber

Antechamber

Chemical shifts prediction:

From structure:

Shiftx2

Sparta+

Camshift

CH3shift- Methyl

ArShift- Aromatic

ShiftS

Proshift

PPM

CheShift-2- Cα

From sequence:

Shifty

Camcoil

Poulsen_rc_CS

Disordered proteins:

MAXOCC

Format conversion & validation:

CCPN

From NMR-STAR 3.1

Validate NMR-STAR 3.1

NMR sample preparation:

Protein disorder:

DisMeta

Protein solubility:

Isotope labeling:

Solid-state NMR:

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07-23-2020, 11:23 PM

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Multiple Site-Specific Phosphorylation of IDPs Monitored by NMR.

Multiple Site-Specific Phosphorylation of IDPs Monitored by NMR.

Related Articles Multiple Site-Specific Phosphorylation of IDPs Monitored by NMR.

Methods Mol Biol. 2020;2141:793-817

Authors: Julien M, Bouguechtouli C, Alik A, Ghouil R, Zinn-Justin S, Theillet FX

Abstract
In line with their high accessibility, disordered proteins are exquisite targets of kinases. Eukaryotic organisms use the so-called intrinsically disordered proteins (IDPs) or intrinsically disordered regions of proteins (IDRs) as molecular switches carrying intracellular information tuned by reversible phosphorylation schemes. Solvent-exposed serines and threonines are abundant in IDPs, and, consistently, kinases often modify disordered regions of proteins at multiple sites. In this context, nuclear magnetic resonance (NMR) spectroscopy provides quantitative, residue-specific information that permits mapping of phosphosites and monitoring of their individual kinetics. Hence, NMR monitoring emerges as an in vitro approach, complementary to mass-spectrometry or immuno-blotting, to characterize IDP phosphorylation comprehensively. Here, we describe in detail generic protocols for carrying out NMR monitoring of IDP phosphorylation, and we provide a number of practical insights that improve handiness and reproducibility of this method.

PMID: 32696390 [PubMed - in process]

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