Simultaneous use of solution NMR and X-ray data in REFMAC5 for joint refinement/detection of structural differences

Area of Science:

• Structural Biology
• Biophysics
• Computational Chemistry

Background:

• X-ray crystallography and Nuclear Magnetic Resonance (NMR) provide complementary structural information.
• Integrating diverse data types can enhance the accuracy and reliability of molecular models.
• Existing refinement programs may not fully leverage the combined power of X-ray and long-range NMR data.

Purpose of the Study:

• To modify the REFMAC5 program for simultaneous refinement using X-ray crystallographic and various NMR data.
• To assess the utility of joint X-ray and NMR refinement for improving structural models.
• To investigate the ability of this integrated approach to detect conformational differences between solid-state and solution states.

Main Methods:

• Modification of the REFMAC5 program (from CCP4) to incorporate pseudocontact shifts, residual dipolar couplings (paramagnetic and diamagnetic).
• Simultaneous refinement of five different protein systems using both X-ray and NMR data.
• Analysis of resulting atomic models for consistency, geometry, and agreement with experimental data.

Main Results:

• Successful integration of X-ray and long-range NMR restraints within REFMAC5.
• Demonstrated increased reliability of atomic models when using complementary data.
• Observed cases where joint refinement yielded a single model consistent with both data types.
• Identified instances revealing distinct protein conformations in solution versus solid-state, beyond experimental uncertainty.

Conclusions:

• The modified REFMAC5 program effectively integrates X-ray and NMR data for structural refinement.
• Joint refinement enhances model reliability and can uncover biologically relevant conformational heterogeneity.
• This approach offers a powerful tool for detailed molecular structure determination and comparison.