A methyl 1 H double quantum CPMG experiment to study protein conformational exchange

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A methyl 1 H double quantum CPMG experiment to study protein conformational exchange

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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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11-25-2018, 06:02 AM

nmrlearner

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A methyl 1 H double quantum CPMG experiment to study protein conformational exchange

A methyl 1 H double quantum CPMG experiment to study protein conformational exchange

Abstract

Protein conformational changes play crucial roles in enabling function. The Carrâ??Purcellâ??Meiboomâ??Gill (CPMG) experiment forms the basis for studying such dynamics when they involve the interconversion between highly populated and sparsely formed states, the latter having lifetimes ranging from ~â??0.5 to ~â??5Â*ms. Among the suite of experiments that have been developed are those that exploit methyl group probes by recording methyl 1H single quantum (Tugarinov and Kay in J Am Chem Soc 129:9514â??9521, 2007) and triple quantum (Yuwen et al. in Angew Chem Int Ed Engl 55:11490â??11494, 2016) relaxation dispersion profiles. Here we build upon these by developing a third experiment in which methyl 1H double quantum coherences evolve during a CPMG relaxation element. By fitting single, double, and triple quantum datasets, akin to recording the single quantum dataset at static magnetic fields of Bo, 2Bo and 3Bo, we show that accurate exchange values can be obtained even in cases where exchange rates exceed 10,000Â*sâ??1. The utility of the double quantum experiment is demonstrated with a pair of cavity mutants of T4 lysozyme (T4L) with ground and excited states interchanged and with exchange rates differing by fourfold (~â??900Â*sâ??1 and ~â??3600Â*sâ??1), as well as with a fast-folding domain where the unfolded state lifetime is ~â??80Â*Âµs.

Source: Journal of Biomolecular NMR

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