NMR paper How to prepare membrane proteins for solid-state NMR: A case study on the alpha-helical integral membrane protein diacylglycerol kinase from E. coli.

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[NMR paper] How to prepare membrane proteins for solid-state NMR: A case study on the alpha-helical integral membrane protein diacylglycerol kinase from E. coli.

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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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How to prepare membrane proteins for solid-state NMR: A case study on the alpha-helical integral membrane protein diacylglycerol kinase from E. coli.

How to prepare membrane proteins for solid-state NMR: A case study on the alpha-helical integral membrane protein diacylglycerol kinase from E. coli.

Related Articles How to prepare membrane proteins for solid-state NMR: A case study on the alpha-helical integral membrane protein diacylglycerol kinase from E. coli.

Chembiochem. 2005 Sep;6(9):1693-700

Authors: Lorch M, Faham S, Kaiser C, Weber I, Mason AJ, Bowie JU, Glaubitz C

Several studies have demonstrated that it is viable to use microcrystalline preparations of water-soluble proteins as samples in solid-state NMR experiments [1-5]. Here, we investigate whether this approach holds any potential for studying water-insoluble systems, namely membrane proteins. For this case study, we have prepared proteoliposomes and small crystals of the alpha-helical membrane-protein diacylglycerol kinase (DGK). Preparations were characterised by 13C- and 15N-cross-polarization magic-angle spinning (CPMAS) NMR. It was found that crystalline samples produce better-resolved spectra than proteoliposomes. This makes them more suitable for structural NMR experiments. However, reconstitution is the method of choice for biophysical studies by solid-state NMR. In addition, we discuss the identification of lipids bound to membrane-protein crystals by 31P-MAS NMR.

PMID: 16138309 [PubMed - indexed for MEDLINE]

Source: PubMed

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