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Protein sample production

NESG target selection DNA cloning protocols Protein expression and purification protocols  Sample Optimization Construct optimization Buffer optimization Cofactor optimization

SDS page gel Protein concentration measurements Assessment of Oligomerization Status gel-filtration and light scattering NMR determined Rotational correlation time Mass spectrum NMR screening

NMR Data Acquisition

Routine operation NMR Sample Preparation Inserting NMR Sample Tuning and matching Deuterium Lock Shimming Pulse width calibration Temperature calibration Chemical shift referencing Advanced operation Deuterium pulse width calibration and decoupling

NMR data acquisition for protein structure determination Common NMR experiment sets Custom NMR experiment setup scripts for VNMRJ Estimation of rotational correlation time Estimation of measurement time Simultaneous 13C,15N-resolved NOESY 2D [13C, 1H]-HSQC for fractionally 13C-labeled samples Long-range 15N-1H correlation experiments for histidine rings Non-uniform sampling (NUS) NUS - Varian NUS - Bruker

Maintenance (VARIAN) Installing and updating BioPack Full probefile calibration Rebooting spectrometer console Conditioning procedure for cryogenic probes

NMR data processing

NMRPipe Brief description of philosophy, commands, and functions 2D experiments 3D 15N and 13C-edited experiments 2D 15N HSQC-TROSY experiment for RDC measurement 2D J-modulation experiment for RDC measurement

Other PROSA TOPSPIN AUTOPROC Processing NUS spectra with MDD NMRPipe processed data conversion to Sparky, CARA, XEASY, and NMRViewJ

NMR Resonance Assignment

• Principles and concepts

Semi-automated protocols CARA Sparky XEASY

Automated resonance assignment AutoAssign AutoAssign server ABACUS PINE server

Validation and deposition Assignment validation suite (AVS) Linear analysis of chemical shift (LACS) Depositing chemical shifts

Structure Calculation and Validation

Principles and concepts

Structure calculation CYANA AutoStructure CS-ROSETTA CS-DP ROSETTA CS-RDC-ROSETTA "Consensus" approaches

Special topics Protein-Ligand complexes Metal ions Residual Dipolar Couplings REDCAT and REDCRAFT Paramagnetic constraints RDC-assisted dimer structure calculation

Structure Refinement CNS refinement XPLOR-NIH refinement ROSETTA refinement

Validation and deposition PdbStat PSVS RPF analysis MolProbity server PDB and BMRB deposition ADIT-NMR HarvestDB SPINS