ATOMIC RESOLUTION STRUCTURE AND CHARGE DENSITY REFINEMENT OF SCORPION ANDROCTONUS AUSTRALIS TOXIN
Farid Benabicha, Virginie Pichon-Pesme, Christian Jelsch & Claude Lecomte 1 , Dominique Housset & Juan Carlos Fontecilla-Camps2
1Laboratoire
de Cristallographie et Modelisation des Matériaux Minéraux et
Biologiques UPRESA CNRS n° 7036, Université Henri
Poincaré, Nancy I - Faculté des sciences, BP 239 - 54506
Vandoeuvres-les-Nancy cedex, Email: lecomte@lcm3b.u-nancy.fr
2Laboratoire
de Cristallographie et Cristallogenese des protéines, Institut
de Biologie Structurale, 41, avenue des Martyrs, 38027
Grenoble Cedex 1, France
Keywords :
electron density, protein, high resolution structure
The TOXIN of Scorpion androctonus Australis Hector is a small protein, which contains 64 residues. It crystallizes in space group P212121 with unit cell parameters a= 45.8 A, b=40.5 A, c= 29.8 A. The diffraction intensities have been measured using a synchroton radiation in Hambourg at 300K. The structure had initially been refined by Dominique Housset [1] using the program SHELXL-93 [2]. There is one molecule per asymmetric unit.
The present work is devoted to study the charge density of this protein, as the resolution is high (d=0.97 A).Due to chemical bonding and atom-atom interaction, the atomic density is not spherical. To describe this non-aspherical character, we use the Hansen-Coppens model [3], in which the electron density in the unit cell is considered as the superposition of the pseudo atomic electron densities. The pseudo atom electron density is given by:
Where ncore and nval are respectively Hartree-Fock spherical core and valence densities. In this aspherical density model, the refined valence population Pval gives an estimation of the net atomic charge q with respect to the number of electron Nval in the free atom valence orbitals: q=Nval-Pval .
The Ylm ,s are
spherical harmonic angular functions of order l in real form, and
Rnl are Slater-type radial functions. Plm
are the multipolar population parameters of each associate
spherical harmonic Ylm, e and e' are the
contraction-expansion coefficients.
The least-squares program MOLLY based on
model [3] was used to determine the atomic coordinates and
thermal parameters. For this protein (1080 atoms), the multipolar
density coefficients are obtained by applying the notion of
transferability [4]. A database of such parameters has been build
in our laboratory from charge density analysis of several peptide
crystals.
In order to assess the quality of the refinement by computation of the Rfree factor, 10% of randomly selected reflections were omitted during the refinement [5].
Three separate refinements I, II, III have been carried out using the MOLLY program .
Refinement IRefinement I
The x,y,z coordinates and Uij temperatures factors were refined with MOLLY using a spherical neutral model.
Refinement IIRefinement II
In this refinement, the multipole Plm parameters from the database [4] were transferred to the Toxin structure. Then the hydrogen atoms were adjusted by moving the H atoms outward along the C-H, N-H and O-H bond directions to bond lengths equal to average values from neutron diffraction studies [6]. The x,y,z coordinates and Uij temperature factors were refined.
Refinement IIIRefinement III
In this refinement the x,y,z coordinates were refined but with application of distance and planarity restrains. The target values of distances were given by F.Allen [6] .
A kappa refinement will be carried out later to evaluate the net atomic charges, because the concept of atomic charge is fundamental to chemistry for understanding the chemical reactivity and physical propreties .
Finally, we will calculate the
electrostatic potentiel because he provides more information than
the electron density mapping about chemical reactivity or
molecule-molecule interaction.
[1] Housset D,.and al.(1994). J. Mol. Biol. 238, 88-103
[2] Sheldrick G.M.,(1993). SHELXL93, Program for crystal structure refinement, University
of Göttigen.Germany
[3] Hansen N.K.& Coppens P. (1978). Acta Cryst., A34, 909-921
[4] Pichon-Pesme V.,Lecomte C.& Lachekar H.(1995).J. Phys. Chem., 99, 6242-6250
[5] Brünger A.T.(1992).Nature (London). 355, 472-474
[6] Allen F.H.(1986).Acta Cryst. B42, 515-522