Erika Schwarz

Small-angle X-ray studies of Lumbricus terrestris haemoglobin

Abstract Small-angle X-ray scattering of Lumbricus terrestris haemoglobin was measured in dilute solutions in 0.1 M Tris HCl buffer, pH 7.0. The following molecular parameters were determined: radius of gyration 11.2 nm, volume 7700 nm3, maximum diameter 29 nm, molecular weight 3.95 × 106. The experimental scattering curve was compared with the scattering curves and distance distribution functions calculated for various models. The overall shape of the haemoglobin could be approximated by a hollow cylinder with the following dimensions: outer radius 13.5 nm, inner radius 5.4 nm, height 16.0 nm. The best fit was obtained with a model which consists of 12 large subunits arranged in two superimposed hexagonal rings with a number of smaller subunits between the large subunits and in the centre of the molecule.

Small-angle X-ray scattering reveals differences between the quaternary structures of oxygenated and deoxygenated tarantula hemocyanin.

Small‐angle X‐ray scattering (SAXS) curves have been recorded for the oxygenated and deoxygenated states of the 4 × 6‐meric hemocyanin from the tarantula Eurypelma californicum. A comparison of the curves shows that the quaternary structures of the two states are different by three criteria, which all indicate that the hemocyanin is less compact in the oxygenated compared to the deoxygenated form: (a) The radius of gyration is 8.65 ± 0.05 nm for the deoxy‐ and 8.80 ± 0.05 nm for the oxy‐form. (b) The maximum particle dimension amounts to 25.0 ± 0.5 nm for the deoxy‐ and to 27.0 ± 0.5 nm for the oxyform. (c) A dip in the intramolecular distance distribution function p(r) is more pronounced and shifted to larger distances in the oxy‐form. The p(r) functions based on SAXS measurements were compared to p(r) functions deduced from published electron microscopical images of three different 4 × 6‐meric hemocyanins from closely related species. The p(r) functions of SAXS and electron microscopy were similar in one case, whereas in the other two cases the distance between the two 12‐meric half‐molecules had to be changed by 1–1.5 nm to obtain good agreement. The differences between the p(r) functions of oxygenated and deoxygenated 4 × 6‐meric tarantula hemocyanin are much larger than one would expect from a comparison of X‐ray structures of the oxygenated and deoxygenated states of a closely related 6‐meric hemocyanin. Thus, the conformational changes upon oxygenation occur at various levels of the quaternary structure, as postulated by hierarchical theories of allosteric interactions.

Small-angle X-ray scattering studies of giant haemoglobin from the annelid Tylorrhynchus heterochaetus

Abstract The extracellular haemoglobin of the polychaete Tylorrhynchus heterochaetus was studied in solution by small-angle X-ray scattering. The following molecular parameters were determined: radius of gyration 10.8±0.2 nm and a maximum intraparticular distance of 29.5±0.5 nm. Models which fit well the experimental data and reflect also the biochemical structure especially the known number of polypeptide chains are presented.

Small-angle X-ray study on the quaternary structure of erythrocruorin from Helisoma trivolvis

Abstract The erythrocruorin from the aquatic snail Helisoma trivolvis was studied in sodium phosphate buffer at pH 6.7 by small angle X-ray scattering. The following molecular parameters were determined: radius of gyration 9.4 ± 0.1 nm and maximum dimension 29 ± 1 nm. A model which fits the experimental data well is presented. The overall shape is best described by a slightly ellipsoidal shape with a hole in the centre. A model consisting of 12 subunits forming a slightly ellipsoidal shape fits very well all scattering data.

Molecular size and shape of the native and reassociated forms of the extracellular haemoglobin of Tubifex tubifex

The extracellular haemoglobin of Tubifex tubifex and the product of its reassociation at neutral pH subsequent to dissociation at alkaline pH, were examined by small-angle X-ray scattering. The following molecular parameters were determined for the native and reassociated molecules, respectively: maximum diameter 30.0±1.0 and 32.0±1.0nm; radius of gyration 10.66±0.15 and 11.07±0.15 nm; molecular weight (3.09±0.15) × 106 and (2.99±0.15) × 106 dalton. Although the scattering curves of the native and reassociated haemoglobin possess similar shapes the distance distribution functions exhibit slight differences in their shape as well as in the position of their maximum. The best fit with the experimental distribution functions was obtained with models consisting of 12 spheres arranged in two hexagonal layers. In the case of the native haemoglobin each of the 12 spheres has a diameter of 9.3 nm while for the reassociated haemoglobin each of the 12 spheres has a diameter of 11.5 nm. The results suggest that although their molecular weights are the same, the reassociated molecule is slightly larger than the native molecule

Small Angle X-Ray Study on the Structure of Active and Inactive Ribulose-1,5-bisphosphate Carboxylase-Oxygenase from Spinach. Evidence for a Configurational Change

Small angle X-ray scattering studies on ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco) from spinach reveal a configurational change in its quaternary structure upon the transition of the molecule from the activated form occurring in the presence of CO2 and Mg2+ to the deactivated form obtained when CO2 and Mg2+ are removed by extensive dialysis under nitrogen. Present structural models are comparable to models which were postulated previously for the same enzyme but isolated from the hydrogen bacterium Alcaligenes eutrophus [O. Meisen- berger, I. Pilz, B. Bowien, G. P. Pal, and W. Saenger, J. Biol. Chem. 259, 4463-4465 (1984)]. The radius of gyration is R = 47.5 ±0.2 nm for the active spinach Rubisco. Upon deactivation, R changes to 49.2 ± 0.2 nm, suggesting a more elongated quaternary structure. The observed difference in deactivation behaviour in ambient and in nitrogen atmosphere indicates a higher affinity of this spinach enzyme to CO2 with respect to the same enzyme from Alcaligenes eutrophus.

X-ray scattering of antibodies: the monomeric 8S subunit of human IgM

Abstract 8S monomeric subunit of a human immunoglobulin M was investigated by small-angle X-ray scattering. The following molecular parameters were determined: radius of gyration 5.9 nm, maximum length 21 nm, hydrated volume 410 nm 3 and two radii of gyration of the cross-section: 2.6 and 1.8 nm. A model equivalent in scattering was found and compared with the model for a human IgG also based on small-angle X-ray scattering data. The Cμ2 domain of the IgM obviously has a very loose structure, and the Fab angle of the 8S IgM (90°) is smaller than that of the IgG (134°).

Small-angle X-ray studies on structure of the subunits of the lac repressor from Escherichia coli

In [l] we reported a small angle X-ray scattering study of the luc repressor and its tetrameric core obtained by proteolytic cleavage, showing both to be highly asymmetric, oblate structures with maximum diameters of 18.0 + 0.5 nm and 16.0 f 0.5 nm, respectively. Here, we report an extension of this study to T-core monomers and dimers, which indicates that the T-core monomers are elongated particles with an axial ratio of 1:2. Dimers are formed by a side-to-side, rather than end-to-end apposition of the 2 monomers and tetramers by an end-to-end apposition of 2 dimers.

Structure of Annelid Hemoglobins — Small Angle X-Ray Studies

Small angle X-ray scattering is a suitable method to get information on the quaternary structure of extracellular hemoglobins in solution. The method allows to calculate directly from the scattering curve and the distance distribution function (p(r)-function) molecular parameters as molecular weight Mr, radius of gyration R and the maximum distance Dmax. The scattering curve reflects the conformation of the macromolecule in the reciprocal space; the distance distribution function reflects the molecule in the real space and represents the frequency of the distances combining any volume element with any other within the molecule. Models describing the conformation of the hemoglobins in solution can be deduced by comparing their scattering curves and p(r)functions with the experimental ones (1, 2).

Small-angle-X-ray study on the quaternary structure of erythrocruorin from Caenestheria inopinata

The erythrocruorin of the clam shrimp Caenestheria inopinata was studied in sodium phosphate buffer at pH 6.8 by small-angle X-ray scattering. The following molecular parameters were determined: radius of gyration 4.77 +/- 0.05 nm, maximum dimension 14.0 +/- 0.5 nm and a volume of 640 +/- 40 nm3. A model which fits the experimental data well is presented. The model is composed of 10 subunits arranged symmetrically in two layers, whereby five subunits are always forming a ring.