NMR paper NMR structural studies of human cystatin C dimers and monomers.

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[NMR paper] NMR structural studies of human cystatin C dimers and monomers.

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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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08-22-2010, 05:08 PM

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NMR structural studies of human cystatin C dimers and monomers.

NMR structural studies of human cystatin C dimers and monomers.

Related Articles NMR structural studies of human cystatin C dimers and monomers.

J Mol Biol. 1997 Aug 15;271(2):266-77

Authors: Ekiel I, Abrahamson M, Fulton DB, Lindahl P, Storer AC, Levadoux W, Lafrance M, Labelle S, Pomerleau Y, Groleau D, LeSauteur L, Gehring K

Human cystatin C undergoes dimerization before unfolding. Dimerization leads to a complete loss of its activity as a cysteine proteinase inhibitor. A similar process of dimerization has been observed in cells, and may be related to the amyloid formation seen for the L68Q variant of the protein. Dimerization is barrier controlled, and no dimer/monomer interconversion can be observed at physiological conditions. As a consequence, very stable, "trapped" dimers can be easily separated from monomers. A study of the structural aspects of cystatin C dimer formation was undertaken using NMR spectroscopy. The monomer/dimer model was verified by (pulse field gradient NMR) self-diffusion molecular mass measurements. Complete backbone resonance assignments and secondary structure determination were obtained for the monomer using data from triple resonance experiments performed on 13C/15N doubly labeled protein. A marked similarity of the cystatin C secondary structure to that of chicken cystatin was observed. Using uniformly and amino-acid-specific 15N-enriched protein, backbone NH signals were assigned for cystatin C in its dimeric state. Comparison of 1H -15N correlation NMR spectra of the monomer and dimer shows that the three-dimensional structure remains unchanged in the dimer and that only local perturbations occur. These are localized to the amino acid residues comprising the cysteine proteinase binding site. Such a mode of dimerization readily explains the complete loss of the inhibitory activity in the dimer. The NMR results also demonstrate that the dimer is symmetric.

PMID: 9268658 [PubMed - indexed for MEDLINE]

Source: PubMed

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