PDBsum entry 1k0y

Oxygen storage/transport

PDB id: 1k0y

Contents

Protein chains

141 a.a. *

146 a.a. *

Ligands

HEM ×4

SO4 ×2

CNO ×2

Waters ×257

* Residue conservation analysis

References listed in PDB file

Key reference

Title

X-Ray crystallographic analyses of symmetrical allosteric effectors of hemoglobin: compounds designed to link primary and secondary binding sites.

Authors

M.K.Safo, T.Boyiri, J.C.Burnett, R.Danso-Danquah, C.M.Moure, G.S.Joshi, D.J.Abraham.

Ref.

Acta Crystallogr D Biol Crystallogr, 2002, 58, 634-644. [DOI no: 10.1107/S0907444902002627]

PubMed id

11914488

Abstract

The rational design and X-ray crystallographic analyses of two symmetrical allosteric effectors of hemoglobin (Hb) are reported. Compound design was directed by the previously solved co-crystal structure of one of the most potent allosteric effectors of Hb, 2-[4-[(3,5-dichlorophenylcarbamoyl)-methyl]-phenoxy]-2-methylpropionic acid (RSR4), which revealed two distinct binding sites for this compound in the Hb central water cavity. The primary binding site has been observed for all compounds of this structural class, which stabilize deoxy Hb by engaging in inter-dimer contacts with three of the four protein subunits. Interactions at the secondary binding site of RSR4 occur primarily between the beta(1) and beta(2) subunits and serve to further constrain the deoxy state. Based on these observations, it was hypothesized that compounds with the ability to simultaneously span and link both of these sites would possess increased potency, but at a lower molar concentration than RSR4. Two symmetrical compounds were designed and synthesized based on this hypothesis. The symmetrical effector approach was taken to minimize the number of compound orientations needed to successfully bind at either of the distinct allosteric sites. X-ray crystallographic analyses of these two effectors in complex with Hb revealed that they successfully spanned the RSR4 primary and secondary binding sites. However, the designed compounds interacted with the secondary binding site in such a way that intra-dimer, as opposed to inter-dimer, interactions were generated. In agreement with these observations, in vitro evaluation of the symmetrical effectors in Hb solution indicated that neither compound possessed the potency of RSR4. A detailed analysis of symmetrical effector-Hb contacts and comparisons with the binding contacts of RSR4 are discussed.

Figure 5.
Figure 5 Stereoviews of allosteric sites of effectors (a) RSR4, (b) TB5-27 and (c) TB5-39. Protein and effector atoms are shown as sticks and structural waters are red spheres. The Hb [1] subunit is light blue, the [2] subunit is magenta, the [1] subunit is orange and the [2 ]subunit is red. C atoms of allosteric effectors are yellow, O atoms are red and N atoms are blue. Dashed black lines indicate hydrogen bonds. For clarity, not all Hb residues lining the allosteric binding sites are shown. Note, for visualization purposes, and owing to secondary-site binding orientation differences between RSR4 and the symmetrical effectors, Hb residue labels and interaction distances for RSR4 in Fig. 5-(a) are labeled on the opposite side of the dimer-dimer interface with respect to labeled residues and interaction distances in the TB5-27-Hb complex (Fig. 5-b) and the TB5-39-Hb complex (Fig. 5-c). For clarity of description, textual reference to RSR4-Hb residue contacts follow dimer-dimer interface residue labeling as pictured for TB5-27 and TB5-39. Importantly, the described RSR4 interactions in the text correspond to equivalent interactions made by this effector at its symmetry-related binding sites and are the mirror image of the contacts labeled in Fig. 5-(a).-.

Figure 6.
Figure 6 Stereoviews of symmetrically related pairs of allosteric effectors bound in the Hb central water cavity. Protein residues are shown as light blue sticks. RSR4 is blue, RSR13 is orange, TB5-27 is yellow and TB5-39 is red. Secondary-site binding for RSR4 is inter-dimer, while an intra-dimer binding mode is adopted by the TB5 compounds.

The above figures are reprinted by permission from the IUCr: Acta Crystallogr D Biol Crystallogr (2002, 58, 634-644) copyright 2002.