Protein and DNA residue orientations in the filamentous virus Pf1 determined by polarized Raman and polarized FTIR spectroscopy.
The Pseudomonas bacteriophage Pf1 is a long ( approximately 2000 nm) and thin ( approximately 6.5 nm) filament consisting of a covalently closed, single-stranded DNA genome of 7349 nucleotides coated by 7350 copies of a 46-residue alpha-helical subunit. The coat subunits are arranged as a superhelix of C(1)()S(5.4)() symmetry (class II). Polarized Raman and polarized FTIR spectroscopy of oriented Pf1 fibers show that the packaged single-stranded DNA genome is ordered specifically with respect to the capsid superhelix. Bases are nonrandomly arranged along the capsid interior, deoxynucleosides are uniformly in the C2'-endo/anti conformation, and the average DNA phosphodioxy group (PO(2)(-)) is oriented so that the line connecting the oxygen atoms (O.O) forms an angle of 71 degrees +/- 5 degrees with the virion axis. Raman and infrared amide band polarizations show that the subunit alpha-helix axis is inclined at an average angle of 16 degrees +/- 4 degrees with respect to the virion axis. The alpha-helical symmetry of the capsid subunit is remarkably rigorous, resulting in splitting of Raman-active helix vibrational modes at 351, 445 and 1026 cm(-)(1) into apparent A-type and E(2)()-type symmetry pairs. The subunit tyrosines (Tyr 25 and Tyr 40) are oriented with phenoxyl rings packed relatively close to parallel to the virion axis. The Tyr 25 and Tyr 40 orientations of Pf1 are surprisingly close to those observed for Tyr 21 and Tyr 24 of the Ff virion (C(5)()S(2)() symmetry, class I), suggesting a preferred tyrosyl side chain conformation in packed alpha-helical subunits, irrespective of capsid symmetry. The polarized Raman spectra also provide information on the orientations of subunit alanine, valine, leucine and isoleucine side chains of the Pf1 virion.
Tsuboi
Masamichi
M
Division of Cell Biology and Biophysics, School of Biological Sciences, University of Missouri-Kansas City, Kansas City, Missouri 64110.
Kubo
Yoshiko
Y
Ikeda
Teruki
T
Overman
Stacy A
SA
Osman
Olivia
O
Thomas
George J
GJ
D000409:Alanine / Q000737:chemistry; D000577:Amides / Q000737:chemistry; D001120:Arginine / Q000737:chemistry; D025561:Bacteriophage Pf1 / Q000737:chemistry* / Q000502:physiology; D002213:Capsid / Q000737:chemistry; D004277:DNA, Single-Stranded / Q000737:chemistry; D004279:DNA, Viral / Q000737:chemistry*; D016679:Genome, Viral; D015591:Microspectrophotometry; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods; D013059:Spectrum Analysis, Raman / Q000379:methods; D014443:Tyrosine / Q000737:chemistry; D014764:Viral Proteins / Q000737:chemistry* / Q000502:physiology; D014771:Virion / Q000737:chemistry / Q000502:physiology; D019065:Virus Assembly / Q000502:physiology
D000577:Amides; D004277:DNA, Single-Stranded; D004279:DNA, Viral; D021122:Protein Subunits; D014764:Viral Proteins; D014443:Tyrosine; D001120:Arginine; D000409:Alanine