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NMR processing:
MDD
NMR assignment:
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MARS
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PINE
Side-chains:
UNIO ATNOS-Ascan
NOEs:
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UNIO Candid
ASDP
Structure from NMR restraints:
Ab initio:
GeNMR
Cyana
XPLOR-NIH
ASDP
UNIO ATNOS-Candid
UNIO Candid
Fragment-based:
BMRB CS-Rosetta
Rosetta-NMR (Robetta)
Template-based:
GeNMR
I-TASSER
Refinement:
Amber
Structure from chemical shifts:
Fragment-based:
WeNMR CS-Rosetta
BMRB CS-Rosetta
Homology-based:
CS23D
Simshift
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:
Shiftcor
UNIO Shiftinspector
LACS
CheckShift
RefDB
NMR model quality:
NOEs, other restraints:
PROSESS
PSVS
RPF scores
iCing
Chemical shifts:
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CheShift2
Vasco
iCing
RDCs:
DC
Anisofit
Pseudocontact shifts:
Anisofit
Protein geomtery:
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PROSESS
What-If
iCing
PSVS
MolProbity
SAVES2 or SAVES4
Vadar
Prosa
ProQ
MetaMQAPII
PSQS
Eval123D
STAN
Ramachandran Plot
Rampage
ERRAT
Verify_3D
Harmony
Quality Control Check
NMR spectrum prediction:
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MestReS
V-NMR
Flexibility from structure:
Backbone S2
Methyl S2
B-factor
Molecular dynamics:
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Amber
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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:
camLILA
ccSOL
Camfold
camGroEL
Zyggregator
Isotope labeling:
UPLABEL
Solid-state NMR:
sedNMR


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Default Expression of human chorionic gonadotropin uniformly labeled with NMR isotopes in Chi

Expression of human chorionic gonadotropin uniformly labeled with NMR isotopes in Chinese hamster ovary cells: an advance toward rapid determination of glycoprotein structures.

Related Articles Expression of human chorionic gonadotropin uniformly labeled with NMR isotopes in Chinese hamster ovary cells: an advance toward rapid determination of glycoprotein structures.

J Biomol NMR. 1996 Jun;7(4):295-304

Authors: Lustbader JW, Birken S, Pollak S, Pound A, Chait BT, Mirza UA, Ramnarain S, Canfield RE, Brown JM

Most secreted eukaryotic proteins are modified by glycosylation, and it has been difficult to solve their structures by crystallographic or NMR techniques because of problems posed by the presence of the carbohydrate. The structure of a chemically deglycosylated form of the human pregnancy hormone, human chorionic gonadotropin (hCG), has been solved by crystallographic methods. Since chemical deglycosylation may have induced changes in the structure, and since it is known that deglycosylated hCG is biologically inactive, the crystallographic structure confirmation by NMR techniques. Also, it has not been possible to determine the structures of the isolated subunits, nor the nature of interactions between the carbohydrate side chains and the protein backbone by crystallographic methods. Structural information via NMR techniques can be obtained from proteins in solution if they can be uniformly labeled with 13C and 15N isotopes. We report the first such uniform labeling of a glycoprotein using a universal 13C- and 15N-labeling medium to express 13C, 15N-labeled hCG, suitable for solving the structure in solution of the native, biologically active form of hCG as well as that of its free subunits. The 13C, 15N-labeled recombinant hCG and its separated subunits are shown to be nearly identical to urinary hCG reference preparations on the basis of protein chemical studies, immunochemistry, biological activity, and the capability of isolated hormone subunits to recombine to form biologically active hormone. Mass spectrometric analysis and preliminary NMR studies indicate that the isotopic labeling is uniform and greater than 90% after only two growth passages in the labeling media. One unexpected finding during subunit purification was that lyophilization of glycoproteins from trifluoroacetic acid HPLC buffers may result in the loss of a significant portion of sialic acid.

PMID: 8765736 [PubMed - indexed for MEDLINE]



Source: PubMed
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