Flow linear dichroism spectra of four filamentous bacteriophages: DNA and coat protein contributions

Abstract

In this study, we have separated the contributions of DNA and protein to the absorption and linear dichroism (LD) of each of four phages: fd, IKe, Pf1, and Pf3. We have found that the DNA packaged in each of the phages is hypochromic relative to the purified single-stranded DNA, suggesting that bases are stacked in all of the phages. We have oriented the phages by flow and for the first time report the intrinsic LD from 320 to 190 nm for each of these phages. From the intrinsic LD of the phages and the isotropic absorption of the individual components, we have determined the reduced dichroism of the DNA within the phages and, subsequently, the maximum angle of inclination of the DNA bases (from the helix axis) for the packaged DNA. The maximum angles were 63 degrees and 64 degrees for the DNAs of class I phages fd and IKe, respectively. The angles were significantly less, 51 degrees and 49 degrees, for the DNAs of the class II phages Pf1 and Pf3, respectively. Thus, the two classes of phage differ in the structures of their packaged DNA, the DNA bases of the class II phages being more parallel to the long axis of the phage than are the DNA bases of the class I phages.

Molecular models and structural comparisons of native and mutant class I filamentous bacteriophages

“…Flow linear dichroism spectra of fd and IKe indicate that the angle between the plane of the Trp [26] side-chain and the virion axis is no more than 43 ° for fd and 41 ° for IKe (Clack & Gray, 1992). To determine tl~is angle for our models, we calculate the direction cosines of the line normal to the leastsquares plane through the Trp ring, and take the angle that this normal makes with the virion axis as the complement of the angle between the plane and the virion axis.…”

The NMR–Rosetta capsid model of M13 bacteriophage reveals a quadrupled hydrophobic packing epitope

“…W26 was thoroughly studied using fiber diffraction (17), UV linear dichroism spectroscopy (47), and different Raman studies (41,46,48). Its sidechain conformation is determined mainly by the two dihedral angles χ 1 (defined by the atoms C'-Cα-Cβ-Cγ) and χ 2 (atoms Cα-Cβ-Cγ-Cδ1; defined as χ 1,2 in some studies).…”

Linear dichroism of visible-region chromophores using M13 bacteriophage as an alignment scaffold

“…Following the conjugation of AF647 to M13 bacteriophage (M13AF647), the LD spectrum of the conjugate was measured, again revealing the UV-region bacteriophage-attributable bands as well as bands attributable to the AF647 cyanine chromophore z -axis 37. As AF647 is too small to flow align, its LD signal in the Couette setup indicates successful conjugation.…”