Supplementary Methods
Raman spectroscopy
Raman spectroscopy was carried out on single crystals of PRD1 also examined by X-ray
diffraction. Each PRD1 crystal, suspended in mother liquor, was sealed within a quartz
capillary of 0.5 mm internal diameter and the Raman spectrum was produced with 785
nm excitation from a solid-state Invictus laser (Kaiser Optical Systems, Inc.) coupled to a
Leica DMLP microscope. The Raman scattering was collected by a microprobe and
transferred via an optical fibre of 100μm diameter to a high-throughput single
monochromator (HoloSpec VPT, Kaiser Optical Systems, Inc.) equipped with a liquidnitrogen cooled CCD (Roper Scientific, model 1340/400). Raman spectra were collected
sequentially from the single crystal and mother liquor. Intense Raman bands from the
mother liquor were removed by subtraction. Spectra were obtained over the region 2001850 cm-1. The spectrum shown in Supplementary Figure 1 represents the average of
approximately 30 exposures of 50 s each.
Estimation of the bulk solvent electron density
The composition of the solvent regions in the crystals was estimated to be 4%(w/v)
Polyethyleneglycol 8000 (PEG 8K), 10% PEG 20K, 0.4M NaCl, 0.1M K2HPO4/KH2PO4
(pH 7.2), based on the compositions of the mother liquor and the doping solution1. A
solution of this composition was measured to have a density of 1.039g/ml and the
corresponding electron density was calculated to be 0.344e-/Å3 by comparison with pure
water, which has an electron density of 0.33e-/Å3 and a density of 0.998g/ml at 293K.
Calculation of the mean number of electrons per lipid
The mass spectrometry data on the viral lipids are summarised in Supplementary Figure
2. The number of electrons N in a lipid molecule with n fatty acid carbon atoms and m
unsaturated (non-consecutive) double bonds is given by
N  8n  2m  c ,
where c is equal to 126, 142 and 234 for PE, PG and CL headgroups respectively. The
mean number of electrons for PE and non-PE type lipids is therefore 390 and 430
respectively. At full hydration, PE headgroups are hydrated by 10 water molecules2 and
PG by 203. Assuming that CL headgroups are hydrated by ~40 water molecules, the mean
number of electrons for PE and non-PE molecules was 490 and 652.
Volumetric analysis of the lipid number
On average, PE and PG molecules have 33 fatty acid carbon atoms with 1.25 and 1.15
double bonds, respectively and CL molecules have 66 fatty acid carbon atoms and 1.68
double bonds. Using these values in conjunction with the current published data on lipid
volumes4, the mean molecular volumes for PE, PG and CL molecules at full hydration
are predicted to be 1450Å3, 1786Å3 and 2903Å3 (the volume of a water molecule being
30.3Å3). Given the lipid composition of the viral membrane, the mean volume per lipid
molecule is 1674Å3. The volume of the membrane estimated from the envelopes for the
two leaflets is 42,402,134Å3 so that the number of lipids in the membrane predicted
volumetrically is 25,330.
References
1.
Cockburn, J. J., Bamford, J. K., Grimes, J. M., Bamford, D. H. & Stuart, D. I.
Crystallization of the membrane-containing bacteriophage PRD1 in quartz
capillaries by vapour diffusion. Acta Crystallogr. D Biol. Crystallogr. 59, 538-40
(2003).
2.
McIntosh, T. J. & Simon, S. A. Area per molecule and distribution of water in
fully hydrated dilauroylphosphatidylethanolamine bilayers. Biochemistry 25,
4948-52 (1986).
3.
Kurze, V., Steinbauer, B., Huber, T. & Beyer, K. A 2-H NMR study of
macroscopically aligned bilayer membranes containing interfacial hydroxyl
residues. Biophys. J. 78, 2441-51 (2000).
4.
Nagle, J. F. & Tristram-Nagle, S. Structure of lipid bilayers. Biochim. Biophys.
Acta 1469, 159-95 (2000).