Modeling of the Dodecameric Subunit of Lumbricus Hemoglobin

Abstract

For modeling the lowresolution shape of the dodecameric subunit of Lumbricus terrestris hemoglobin, experimental smallangle X-ray scattering (SAXS) data and ab initio modeling approaches using a genetic algorithm or simulated annealing have been applied. In addition to the use of strict ab initio approaches, procedures which additionally include available structural information concerning symmetry and shape in the form of constraints or templates have been employed to improve the results. Templates for the subunit were preferably derived from SAXS-based models for the native hexagonal bilayer (HBL) complex that were biased by electron microscopic reconstructions. The obtained subunit models were carefully examined by variation of different selection and averaging methods and other checks such as surface renderings of the models. The findings were quantified by prediction of scattering profiles, I(h) and p(r), and structural and hydrodynamic parameters (V , R G , d max , s, D). The best matching models for the subunit were also scrutinized by comparing them to a model derived from currently available crystallographic data. The following results could be obtained: (i) The obtained parameter predictions for the dodecameric subunit are satisfactory, if compared to the SAXS data (consensus model, profiles and molecular parameters) or the results from hydrodynamic studies. (ii) The comparison between solution and crystal data of the dodecameric subunit, however, unequivocally proves a different behavior of the subunit in solution and the crystalline state.

A comparison of protein models obtained by small-angle X-ray scattering and crystallography

“…In contrast to previous findings (Zipper et al, 2006), owing to an improved set of experimental data, the comparison of the observed and predicted scattering behaviour of the dodecameric subunit of L. terrestris Hb does not hint at any obvious discrepancies between the shapes in solution and the crystal. The previously supposed discrepancies could be traced back to a hitherto overlooked deficiency in the desmeared experimental scattering curve, which led to underestimated particle dimensions in the previous studies (Krebs, Kuchumov et al, 1996;Zipper et al, 2006) as compared with the revised data in Table 1. As may be taken from the SAXS profiles ( Fig.…”

“…Some deficiencies in the previous desmearing procedure also necessitated a scrupulous re-evaluation of the initial SAXS data. The data in this paper are slightly different from the previous data (Krebs, Kuchumov et al, 1996), and recently supposed discrepancies between solution and crystal data (Zipper et al, 2006) have now been removed.…”

“…In all cases SAXS and hydrodynamic information is available in addition to the availability of the aminoacid (AA) sequence (SWISS-PROT: Boeckmann et al, 2003) and crystallographic data [Protein Data Bank (PDB): Berman et al, 2000]. The most important features and references of the analyzed proteins are given below (for further details see Zipper et al, 2005Zipper et al, , 2006Durchschlag et al, 2006).…”

“…In general, experimental data are taken from the references cited in x2. Details of modelling procedures are given in previous papers (Zipper et al, 2005(Zipper et al, , 2006Durchschlag et al, 2006). In the case of SAXS-based modelling, if applicable, background subtraction (BC) was used.…”