Robert M. Cooke

The solution structures of epidermal growth factor and transforming growth factor alpha.

The structures of human epidermal growth factor (EGF) and human transforming growth factor alpha (TGF alpha) have been determined in solution using nuclear magnetic resonance techniques. The features of each structure are described and similarities and differences between them are discussed. The structures are combined with information from sequence homologies to produce a model of the receptor-recognition sites of EGF and TGF alpha, which can be tested in a site-directed mutagenesis programme. The model assists in explaining previous observations of sequence-activity relationships. The TGF alpha and EGF structures also serve as models for homologous modules in other extracellular proteins.

Structure-function analysis of epidermal growth factor: site directed mutagenesis and nuclear magnetic resonance

The role of leucine‐47 in determining the structure and activity of human epidermal growth factor was examined using site‐directed mutagenesis. Wild type protein and four variants in which Leu47 was replaced by valine, glutamate, aspartate and alanine were produced from yeast. 1H NMR experiments demonstrated that substitution of Leu47 had little effect on the protein structure. The observed reduction in receptor binding affinity caused by the substitutions could thus be attributed to perturbation of a residue directly involved in receptor interactions.

A high resolution 1H NMR study of the solution structure of human epidermal growth factor

500 MHz 1H NMR studies of human epidermal growth factor are described. The backbone resonances of the 1–48 derivative of hEGF have been assigned using two‐dimensional techniques. Analysis of the type and magnitude of the observed sequential nuclear Overhauser effects and the NH‐αCH spin‐spin coupling constants allowed prediction of the secondary structure. Aspects of the tertiary structure are also identified. A pair of antiparallel β‐sheets involving residues 18–23 and 28–34 is a dominant feature of the solution structure.

A 1H NMR study of the solution conformation of the neuropeptide galanin

The conformations of the neuropeptide galanin in water and trifluoroethanol solutions have been examined by 1H NMR spectroscopy. Analysis of two-dimensional NMR experiments enabled the assignment of virtually all the 1H resonances of galanin in trifluoroethanol solution and many of the 1H resonances in aqueous solution. Interpretation of the NMR data in structural terms suggests that in trifluoroethanol galanin is predominantly helical while in water it does not adopt a fixed conformation.

Structure-function relationships in epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-α)

Abstract The solution structures of the homologous growth factors human epidermal growth factor (hEGF) and human transforming growth factor-alpha (hTGF-α), as determined by high resolution NMR and various computational methods, are described. Knowledge of these structures and the sequences of other homologous proteins leads to predictions about growth factor residues which may be involved in the receptor/ligand interface. Recent experiments designed to check these predictions are described briefly. These involve site-specific mutagenesis, receptor binding assays and high resolution NMR studies.

High resolution 1H NMR study of the solution structure of the S4 segment of the sodium channel protein

Na‐ channel; Peptide, S4‐; NMR; Peptide structure

Structure function relationships in EGF, TGF-α and IGFI

Summary The solution structures of the homologous growth factors hEGF and hTGF-α, have been determined independently from high resolution nuclear magnetic resonance (NMR) data. A model of the insulin-like growth factor structure based on insulin coordinates (Blundell et al. (1978) Proc natn. Acad. Sci. U.S.A.75, 180–184), has also been refined using molecular dynamics simulations with NMR-determined restraints. Knowledge of these structures, together with known sequences of other homologous proteins and experiments with site-specific residue changes, allows predictions to be made about growth factor residues which might be involved in the receptor–ligand interfaces.