Small-angle X-ray studies of the quaternary structure of the lac repressor from Escherichia coli.

The quarternary structures of the lac repressor molecule from Escherichia coli and its tetrametic core, which can be derived from it by proteolytic cleavage, were studied in dilute solutions by small-angle X-ray scattering. The dimensions and general shape of the lac repressor and of the tetrameric core are reported. The core itself appears to be an elongated structure, and in the intact repressor the headpieces are located at its ends. The results ar derived from model calculations and from the following molecular parameters determined from the scattering curve and the pair distance distribution function: for lac repressor, radius of gyration 5.30 +/- 0.02 nm, radius of gyration of the cross section 2.20 +/- 0.03 nm, maximum diameter 18.0 +/- 0.5 nm, hydrated volume 329 +/- 20 nm3, relative molecular mass 149 000 +/- 15 000, for tetrameric core, radius of gyration 4.92 +/- 0.02 nm, radius of gyration of the cross section 2.24 +/- 0.03 nm, maximum diameter 16.0 +/- 0.5 nm, hydrated volume 278 +/- 15 nm3, relative molecular mass 120 000 +/- 10 000.

[Small-angle X-ray-scattering investigation and structural-model study of the fatty-acid synthetase from pig liver (author's transl)]. / Röntgenkleinwinkel-Untersuchungen und Strukturmodell-Betrachtungen an der Fettsäuresynthetase aus Schweineleber.

The structure of the fatty acid synthetase from pig liver was studied on models based upon structural and functional properties selected from pertinent results available from numerous investigations carried out with fatty acid synthetases from this and other sources. When comparing small-angle X-ray-scattering curves calculated with these models and curves obtained from small-angle X-ray-scattering experiments carried out with the pig-liver enzyme, we tried to select a model which would lead to an acceptable correlation between the calculated and the experimental curves and at the same time fulfil the known structural and functional requirements. The comparison of the curves was started with a model of low complexity. The observed discrepancy, together with arguments from the structural and the functional properties, helped decide which is the next most reasonable model to be considered. This procedure was repeated for five models of increasing complexity. In the model which led to the best fit the multienzyme complex is composed of two halves in an assymetric conformation including hollow spaces. This highly anisotropic model would imply that the two halves change their conformation each time a synthetic cycle is completed and that the growing fatty acid is handed over from one half to the other.

The lipid bilayer structure of the abnormal human plasma lipoprotein X. An X-ray small-angle-scattering study.

The structure of the abnormal lipoprotein X occurring in the plasma of patients with obstructive jaundice was investigated by X-ray small-angle scattering. The data were analyzed by discussing the distance distribution functions obtained directly from the experimental data by Fourier transformation, involving no a priori assumptions. The results provide evidence for lipoprotein X being essentially a random distribution of lamellae with a thickness of 5.1 nm and are consistent with hollow spherical (vesicular) structures of outer diameters greater than 30 nm with some overall size heterogeneity. Under the experimental conditions chosen, lipoprotein concentrations between 0.01 and 0.18 g/ml in in buffers of low ionic strength, lateral stacking as observed in negative-stain electron microscopy does not occur. The electron density profile perpendicular to the lamellar plane indicates that a lipid bilayer is the underlying structural principle, with the protein moieties partly bound within the polar head-group regions and partly occluded in soluble form in the vesicle interior.

On the conformation of serine-specific transfer RNA. Studies by small-angle X-ray scattering and ultraviolet absorption of the molecule in solution.

The scattered X-ray intensities from dilute solutions of tRNASer (yeast) in 0.1 M Soerensen buffer at pH 7.0 were measured at 25 degrees C. The radius of gyration, molecular weight and volume were determined. A model equivalent in scattering is given. The change of the conformation of tRNASer by heating was followed by small-angle X-ray measurements and ultraviolet absorption in a temperature range 20-70 degrees C. The molecule begins to unfold at about 40 degrees C and 70 degrees C has a random coil conformation. Addition of magnesium stabilizes the tRNASer molecule. The reversibility of the melting process was also studied by both methods. An interesting effect was found by ultraviolet absorption: by heating the tRNASer solutions to 55 degrees C and 60 degrees C and subsequently slowly cooling, the melting curves lie at higher absorption values than the corresponding cooling curves. The small-angle data and optical properties of tRNASer are compared with those of tRNAPhe which has already been thoroughly investigated.

Small-angle x-ray studies on the structure of 16-S ribosomal RNA and of a complex of ribosomal protein S4 and 16-S ribosomal RNA from Escherichia coli.

16-S ribosomal RNA and a complex of ribosomal protein S4 and 16-S rRNA were studied in solution by small-angle X-ray scattering. Concentration series of the 16-S rRNA and the S4 - 16-S-rRNA complex were measured in 37.5 mM Tris-HCl buffer pH 7.4 at 5 degrees C. The following data were determined. The radii of gyration for the 16-S rRNA and S4 - 16-S-rRNA complex were R = 17.6 +/- 0.6 nm, respectively. The two respective values of the radii of gyration of the cross-section were Rq,1 = 8.42 +/- 0.1 nm and 8.33 +/- 0.3 nm, and Rq,2 = 0.988 +/- 0.03 nm and 0.996 +/- 0.03 nm. The largest diameters of the 16-S RNA and S4 - 16-S-RNA complex were L = 61.8 +/- 1 nm and 60.0 +/- 1 nm, respectively. Volumes of V = 1570 +/- 60 nm3 were found for both particles. In the Tris buffer used, no significant differences were found between the scattering curves of 16-S rRNA and the complex is a flat elliptical cylinder with the following dimensions: large axis 61.7 nm, small axis 35.4 nm and height 2 nm. The theoretical scattering curve fits the experimental one as long as the shape of the measured curve is due only to the overall shape of the particle. A model equivalent over the whole measured angular range is one built up from a large number of spheres that simulate the known substructure of the RNA. The outer dimensions of this model correspond to those of the flat elliptical cylinder.

Shape and volume of fragments Fab' and (Fab')2 of anti-poly(D-alanyl) antibodies in the presence and absence of tetra-D-alanine as determined by small-angle x-ray scattering.

The conformation of two fragments derived from anti-poly(D-alanyl) antibodies, the divalent fragment (Fab')2 and the monovalent fragment Fab', was studied by small-angle X-ray scattering before and after interaction with the tetra-D-alanine amide hapten. More than 90% of the combining sites were occupied by the hapten. No significant changes were observed in the volume or in the radius of gyration, with either of the fragments. This contrasts with the significant decrease in these two parameters found upon reacting the hapten with intact anti-poly(D-alanyl) antibodies (I. Pilz, O. Kratky, A. Licht, and M. Sela (1973), Biochemistry 12, 4998). For Fab', the radius of the whole particle was found to be 3.48 nm in the absence of the hapten and 3.46 nm in its presence, the radius of gyration of the cross-section was 1.37 nm without hapten and 1.38 nm in its presence, and the volume of the particle was 98 nm3 in the absence of the hapten and 91 nm3 in its presence. For (Fab')2 the respective values were 5.06 and 5.05, 1.38 and 1.37, and 182 and 182. These results suggest that a conformational change occurs within the antibody molecule, but not within its Fab fragment, upon reaction with the tetraalanine hapten.