We have found a correlation between liposome fusion kinetics and lipid phase behavior for several inverted phase forming lipids. N-Methylated dioleoylphosphatidylethanolamine (DOPE-Me), or mixtures of dioleoylphosphatidylethanolamine (DOPE) and dioleoylphosphatidylcholine (DOPC), will form an inverted hexagonal phase (HII) at high temperatures (above TH), a lamellar phase (L alpha) at low temperatures, and an isotropic/inverted cubic phase at intermediate temperatures, which is defined by the appearance of narrow isotropic 31P NMR resonances. The phase behavior has been verified by using high-sensitivity DSC, 31P NMR, freeze-fracture electron microscopy, and X-ray diffraction. The temperature range over which the narrow isotropic resonances occur is defined as delta TI, and the range ends at TH. Extruded liposomes (approximately 0.2 microns in diameter) composed of these lipids show fusion and leakage kinetics which are strongly correlated with the temperatures of these phase transitions. At temperatures below delta TI, where the lipid phase is L alpha, there is little or no fusion, i.e., mixing of aqueous contents, or leakage. However, as the temperature reaches delta TI, there is a rapid increase in both fusion and leakage rates. At temperatures above TH, the liposomes show aggregation-dependent lysis, as the rapid formation of HII phase precursors disrupts the membranes. We show that the correspondence between the fusion and leakage kinetics and the observed phase behavior is easily rationalized in terms of a recent kinetic theory of L alpha/inverted phase transitions. In particular, it is likely that membrane fusion and the L alpha/inverted cubic phase transition proceed via a common set of intermembrane intermediates.

Membrane fusion and inverted phases.

Solution structure of human and mouse immunoglobulin M by synchrotron X-ray scattering and molecular graphics modelling. A possible mechanism for complement activation.

A new x‐ray beamline has recently been built, and is now operational, on dipole magnet 2 of the Synchrotron Radiation Source (SRS) at Daresbury. This beamline takes 32 mrad of horizontal aperture from a central tangent point. The time‐resolved x‐ray diffraction (TRXD) Station 2.1, takes 17 mrad of horizontal aperture and is the central point of this paper. Beamline 2 has been realized as part of a SERC–MRC agreement.

Time‐resolved x‐ray diffraction station: X‐ray optics, detectors, and data acquisition

Pentameric and decameric structures in solution of serum amyloid P component by X-ray and neutron scattering and molecular modelling analyses.

Factor H is a regulatory component of the complement system. It has a monomer Mr of 150,000. Primary structure analysis shows that the polypeptide is divided into 20 homologous regions, each 60 amino acid residues long. These are independently folding domains and are termed "short consensus repeats" (SCRs) or "complement control protein" (CCP) repeats. High-flux synchrotron X-ray and neutron scattering studies were performed in order to define its solution structure in conditions close to physiological. The Mr of factor H was determined as 250,000-320,000 to show that factor H is dimeric. This structure is maintained at concentrations between 1 and 11 mg/mL in the pH range 5-9. Zn2+ ions are an inhibitor of C3b cleavage by factor I, a reaction in which factor H acts as a cofactor. Additions of Zn2+ to factor H caused it to form oligomers containing 4-10 monomers. The radius of gyration RG of native factor H by X-rays or by neutrons in 0% or 100% 2H2O buffers is not measurable but is greater than 12.5 nm. Two cross-sectional radii of gyration RXS-1 and RXS-2 were determined as 3.0-3.1 and 1.8 nm, respectively. Analyses of the cross-sectional intensities show that factor H is composed of two distinct subunits. The RXS-1 corresponds to the cross-sectional properties of both subunits and exhibits an unusual radiation dependence on the X-ray flux. Since RXS-2 is close to the corresponding RXS of C4b binding protein (91% of which is formed from SCR/CCP domains), it is inferred that the SCR/CCP domains of factor H and C4b binding protein have similar solution structures. The use of hydrodynamic spheres to reproduce literature sedimentation coefficients of 5.5-5.6 S showed that these were compatible with a V-shaped arrangement of two rods (36 spheres each, length 87 +/- 5 nm) joined at an angle of 5 degrees. The use of a similar arrangement of 244 spheres arranged in two rods (length 77 nm) to fit the experimental X-ray and neutron scattering curves showed that the two rods are joined at an angle of 5 degrees. This model corresponds to an actual RG of 21-23 nm. The separation between each SCR/CCP in factor H is close to 4 nm. In the solution structure of factor H, the SCR/CCP domains are in a highly extended conformation.

Oligomeric domain structure of human complement factor H by X-ray and neutron solution scattering.

The small-angle scattering facility at Daresbury has been constructed for diffraction studies of a wide range of naturally occurring and synthetic materials The high brightness of the SRS is combined with focusing optics, resulting in exposure times that can be two or three orders of magnitude less than those required on a conventional source Spacings of 2000 A in the vertical direction and 300 A in the horizontal direction can be observed, while the resolution between diffraction orders is 5000 and 600 A In addition, preliminary results have been obtained on a double-crystal diffractometer that has a resolution, in one dimension, of better than 26 000 A For high-angle fibre diffraction studies, a camera with pinhole collimation has been constructed Examples from solution scattering and fibre diffraction are used to illustrate the performance of these facilities

A. W. Thompson

Papers

Facilities for solution scattering and fibre diffraction at the Daresbury SRS