Sequential 1H and 15N NMR resonance assignment and secondary structure of ferrocytochrome c2 from Rhodobacter sphaeroides.
Sequence-specific 1H and 15N assignments have been made for the amino acids of the ferrocytochrome c2 from Rhodobacter sphaeroides. Initial assignments were made by analysis of a series of homonuclear 2D COSY, TOCSY, and NOESY spectra obtained with the unlabeled protein. 2D and 3D 1H-15N correlated spectra obtained for a uniformly 15N-labeled ferrocytochrome c2 were used to confirm and extend the assignments. Partial 13C assignments have also been made by means of HSQC experiments on 13C at natural abundance, in particular for about two-thirds of the 13C alpha. Medium-range NOE connectivities, together with 3J(HC alpha NH) coupling constants, indicated the presence of five helices at positions 6-16, 60-68, 74-82, 84-91, and 109-120. No other regular secondary structure was observed. This folding is similar to that previously observed for the ferrocytochrome c2 of Rhodobacter capsulatus in solution, which exhibits approximately 50% sequence identity. Moreover, the rotation rates of the aromatic rings of phenylalanine or tyrosine, when conserved, were similar to those observed for R. capsulatus. Furthermore, C alpha H chemical shifts, which are sensitive to the secondary structure and ring current effects of the heme, appear to be very similar for the two proteins. Consequently, the solution structure of R. sphaeroides ferrocytochrome c2 appears to be very similar to that of R. capsulatus ferrocytochrome c2. These results are compared with the X-ray crystal structure of the R. sphaeroides ferrocytochrome c2.
Gans
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Institut de Biologie Structurale CEA-CNRS Jean-Pierre Ebel, Grenoble, France. pierre@rmn.ibs.fr
Simorre
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JP
Caffrey
M
M
Marion
D
D
Richaud
P
P
Verméglio
A
A
D000595:Amino Acid Sequence; D002247:Carbon Isotopes; D003574:Cytochrome c Group / Q000737:chemistry*; D045362:Cytochromes c2; D006418:Heme / Q000737:chemistry; D006859:Hydrogen; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009587:Nitrogen Isotopes; D017433:Protein Structure, Secondary; D016354:Rhodobacter capsulatus / Q000378:metabolism; D012242:Rhodobacter sphaeroides / Q000378:metabolism*
D002247:Carbon Isotopes; D003574:Cytochrome c Group; D009587:Nitrogen Isotopes; D006418:Heme; D006859:Hydrogen; D045362:Cytochromes c2