A large portion of the 13C resonance assignments for murine epidermal growth factor (mEGF) at pH 3.1 and 28°C has been determined at natural isotope abundance. Sequence-specific 13C assignments are reported for 100% of the assignable Cα, 96% of the Cβ, 86% of the aromatic and 70% of the remaining peripheral aliphatic resonances of mEGF. A good correlation was observed between experimental and back-calculated Cα chemical shifts for regions of regular β-sheet structure. These assignments also provide the basis for interpreting 1Hα-13Cα heteronuclear NOE (HNOE) values in mEGF at natural isotope abundance. Some of the backbone polypeptide segments with high internal mobility, indicated by these 1Hα-13Cα HNOE measurements, correlate with locations of residues involved in the putative mEGF-receptor binding site. Using four families of mEGF structures obtained over the last few years, we demonstrate that standard deviations between experimental and back-calculated ΔδCα values can be used to monitor the refinement of this protein's structure, particularly for β-sheet regions. Improved agreement between calculated and observed values of ΔδCα is correlated with other measures of structure quality, including lowered values of residual constraint violations and more negative values of conformational energy. These results support the view that experimental conformation-dependent chemical shifts, ΔδCα, can provide a reliable source of information for monitoring the process of protein structure refinement and are potentially useful restraints for driving the refinement.
All Science Journal Classification (ASJC) codes
- Epidermal growth factor
- Molecular dynamics
- Nuclear relaxation
- Pulsed-field gradients