Proton NMR studies of transforming and nontransforming H-ras p21 mutants.

摘要: One- and two-dimensional nuclear magnetic resonance spectroscopy (1D 2D NMR) site-directed mutagenesis were used to study the influence of mutations on conformation H-ras oncogene product p21. No severe structural differences between different mutants, whether they transforming or nontransforming, could be detected. Initially, selective incorporation 3,5-deuterated tyrosyl residues into p21 NMR identify resonances representing spin system imidazole rings three histidyl in protein, six nine rings, four five phenylalanyl rings. The systems phenyl Phe{sup 28}, 78}, 82} assigned by using mutant proteins, since no structure-induced spectral changes aromatic part spectra proteins Sequence-specific assignments histidine obtained comparison distance information Overhauser enhancement (NOESY) experiments with crystal structure. change chemical shift values H1{prime} proton {alpha}-phosphate bound GDP (F28L) 28-fold increase GDPmore » dissociation rate constants this suggest a strong interaction 28} p21-bound nucleotide. In solution, GDP{center dot}Mg{sup 2+} has an anti conformation, ring is close ribose 2+}.« less