Modeling techniques for analysing conformational transitions in hemocyanins by small-angle scattering of X-rays and neutrons

Structural and Conformational Analysis of Scorpion (Buthus sindicus) Hemocyanin Using Low Resolution Techniques

“…cally related scorpion A. australis hemocyanin calculated from electron microscopy as 92.2 Å has recently been corrected to 88.5 Å [27,29]. Unfortunately, we could not manage yet the comparison in terms of shape calculation of the oxygenated/deoxygenated states of scorpion native hemocyanin from the SAXS data alone (also not reported for any other hemocyanin due to the enormous size of the molecule) but our EM studies and identical Rg values confirm the previous hypothesis that hemocyanin become less compact upon oxygenation and there is a sharp conformational switch in terms of flexible tilting in the two identical 2x (2x6-mers) half-molecules against each other in the native 4x6-mers [24][25][26][28][29]. The reasons for this discrepancy be- 86.5 Å ± 0.5) confirm the previous data suggesting that the oxygenated hemocyanin is longer than the deoxygenated hemocyanin by almost 2 Å [28][29].…”

“…2f. These observations in AFM are also complemented by the various independent studies on 4x6-mers hemocyanins under oxygenated and deoxygenated states examined by other low resolution techniques such as electron microscopy and/or solution X-ray scattering [25][26][27][28][29]. 2h and j also revealed some undefined molecular orientations which probably belongs to the lateral views of the native molecule.…”

“…On the other hand, three-dimensional structure of the hemocyanin subunit II from the closely related horseshoe crab (L. polyphemus) is known at atomic resolution in both oxygenated/deoxygenated states (2.18 Å and 2.4 Å, respectively) and reveal almost no differences within rms of 0.3 Å [33][34]. distance (~2.5 Å) between two trimers of each hexamer, rotation of the hexamers (~26°) within 2x6-mers against each other and the angle (13°) between the two identical 2x6-mers as well as a 14 Å shift (tilt) in the 2x6-meric half-molecule with respect to the rest identical half within the native 4x6-mers in tarantula hemocyanin during breathing [25][26][28][29]. It is also known from various studies that a single subunit (for e.g.…”

“…4A) has been calculated from the scattering profile alone assuming a two-fold symmetry which is not exactly true but acceptable for the resolution (L=6) used for the shape restoration [22][23]. 2) as well as previous independent studies by various authors [24][25][26]28]. Moreover, molecular envelope calculated for the native oxygenated hemocyanin also depicts an apparent slight tilt of the 2x6mers with respect to the rest of half molecule within the native 2x (2x6-mers) assembly and nicely complement our EM results (Fig.…”

ELLSTAT: shape modeling for solution small-angle scattering of proteins and protein complexes with automated statistical characterization

“…There are an increasing number of methods for producing low-resolution models of proteins and protein complexes in solution from small-angle scattering data. Ab initio shaperestoration methods (Chacó n et al, 1998;Svergun, 1999;Walther et al, 2000;Svergun et al, 2001;Heller et al, 2003;Hartmann et al, 2004), in particular, have become increasingly popular as biologists apply solution scattering methods to proteins and protein complexes that are either difficult to crystallize or too large for high-resolution structure determination by NMR. In many cases, significantly simplified models can still play an important role in interpreting small-angle scattering data of proteins and protein complexes.…”