Amide modes of the alpha-helix: Raman spectroscopy of filamentous virus fd containing peptide 13C and 2H labels in coat protein subunits.
The filamentous virus fd consists of a single-stranded DNA genome sheathed by 2700 copies of a 50-residue alpha-helical subunit (protein pVIII) and serves as a model assembly of alpha-helices. To advance vibrational assignments for the alpha-helix, we have investigated Raman spectra of fd virions containing 13C and 2H (deuterium) labels at various main-chain sites of the pVIII subunits. 13C was introduced at specific peptide carbonyls, while deuterium was introduced at selected alpha-carbon (Calpha) and amide nitrogen positions. Interpretation of the Raman spectra reveals a previously unrecognized alpha-helix band in the spectral interval 730-745 cm-1, tentatively assigned to a carbonyl in-plane bending mode (amide IV). Experimental evidence has also been obtained for a distinctive alpha-helix marker near 1345 cm-1, assigned to a coupled Calpha-H bending and Calpha-C stretching mode. The fd virions containing 13C-labeled carbonyls exhibit unexpectedly complex amide I profiles, consisting of multiple band components. Amide I splitting resulting from 13C substitution of carbonyls is attributed to decoupling of transition-dipole interactions normally occurring in the extended pVIII helix. The present study identifies novel conformation-dependent Raman bands in a native alpha-helix assembly, confirms amide I and amide III assignments proposed previously for filamentous viruses, and facilitates new Raman assignments for the packaged ssDNA. The alpha-helix markers identified here should also be useful in conformation analyses of other proteins by Raman spectroscopy.
Overman
S A
SA
Division of Cell Biology and Biophysics, School of Biological Sciences, University of Missouri-Kansas City 64110, USA.
Thomas
G J
GJ
D000577:Amides; D002213:Capsid / Q000737:chemistry* / Q000378:metabolism; D002247:Carbon Isotopes; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism; D003903:Deuterium; D017912:Inovirus / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D017433:Protein Structure, Secondary; D013059:Spectrum Analysis, Raman; D019065:Virus Assembly
D000577:Amides; D002247:Carbon Isotopes; D004277:DNA, Single-Stranded; D004279:DNA, Viral; D010455:Peptides; D003903:Deuterium