COMPLETE C-13 ASSIGNMENTS FOR RECOMBINANT CU(I) RUSTICYANIN PREDICTION OF SECONDARY STRUCTURE FROM PATTERNS OF CHEMICAL-SHIFTS

Complete resonance assignments for the C-13 spectrum of reduced (Cu(I)) rusticyanin have been made using C-13,N-15 doubly labeled recombinant material, The reported assignments include those for the carboxyl and carbonyl carbon atoms and protonated aromatic ring carbons, and were obtained using a variety of 2- and 3D inverse-detected NMR experiments, including C-13,N-15,H-1 triple resonance experiments and HCCH-COSY and -TOCSY, Backbone carbonyl assignments were obtained using 3D HNCO and HCACO spectra, and modified versions of 2D H(CA)CO and HMBC spectra were used to obtain side-chain carboxgl carbon and methionine epsilon-methyl carbon assignments, respectively, A comparison of the C-13(alpha), C-13(beta) and (CO)-C-13 chemical shifts with published 'random coil' values confirms the conclusion reached from a consideration of the (3)J(HN alpha) coupling constants and the pattern of sequential NOEs, that the protein consists largely of beta-structure.