PDBsum entry 3vhb

Oxygen storage/transport

PDB id: 3vhb

Contents

Protein chains

135 a.a. *

Ligands

HEM-IMD ×2

Waters ×87

* Residue conservation analysis

PDB id:

3vhb

Name:

Oxygen storage/transport

Title:

Imidazole adduct of the bacterial hemoglobin from vitreoscilla sp.

Structure:

Protein (hemoglobin). Chain: a, b. Synonym: soluble cytochrome o. Engineered: yes

Source:

Vitreoscilla stercoraria. Organism_taxid: 61. Strain: c1. Atcc: atcc 15218. Collection: atcc 15218. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.10Å R-factor: 0.238

Authors:

M.Bolognesi,A.Boffi,M.Coletta,A.Mozzarelli,A.Pesce,C.Tarricone, P.Ascenzi

Key ref:

M.Bolognesi et al. (1999). Anticooperative ligand binding properties of recombinant ferric Vitreoscilla homodimeric hemoglobin: a thermodynamic, kinetic and X-ray crystallographic study. J Mol Biol, 291, 637-650. PubMed id: 10448042 DOI: 10.1006/jmbi.1999.2975

Date:

17-Mar-99 Release date: 18-Aug-99

Protein chains

P04252  (BAHG_VITST) -  Bacterial hemoglobin from Vitreoscilla stercoraria

Seq: 146 a.a.

Struc: 135 a.a.

DOI no: 10.1006/jmbi.1999.2975

J Mol Biol 291:637-650 (1999)

PubMed id: 10448042

Anticooperative ligand binding properties of recombinant ferric Vitreoscilla homodimeric hemoglobin: a thermodynamic, kinetic and X-ray crystallographic study.

M.Bolognesi, A.Boffi, M.Coletta, A.Mozzarelli, A.Pesce, C.Tarricone, P.Ascenzi.

ABSTRACT

Thermodynamics and kinetics for cyanide, azide, thiocyanate and imidazole binding to recombinant ferric Vitreoscilla sp. homodimeric hemoglobin (Vitreoscilla Hb) have been determined at pH 6.4 and 7.0, and 20.0 degrees C, in solution and in the crystalline state. Moreover, the three-dimensional structures of the diligated thiocyanate and imidazole derivatives of recombinant ferric Vitreoscilla Hb have been determined by X-ray crystallography at 1.8 A (Rfactor=19.9%) and 2.1 A (Rfactor=23.8%) resolution, respectively. Ferric Vitreoscilla Hb displays an anticooperative ligand binding behaviour in solution. This very unusual feature can only be accounted for by assuming ligand-linked conformational changes in the monoligated species, which lead to the observed 300-fold decrease in the affinity of cyanide, azide, thiocyanate and imidazole for the monoligated ferric Vitreoscilla Hb with respect to that of the fully unligated homodimer. In the crystalline state, thermodynamics for azide and imidazole binding to ferric Vitreoscilla Hb may be described as a simple process with an overall ligand affinity for the homodimer corresponding to that for diligation in solution. These data suggest that the ligand-free homodimer, observed in the crystalline state, is constrained in a low affinity conformation whose ligand binding properties closely resemble those of the monoligated species in solution. From the kinetic viewpoint, anticooperativity is reflected by the 300-fold decrease of the second-order rate constant for cyanide and imidazole binding to the monoligated ferric Vitreoscilla Hb with respect to that for ligand association to the ligand-free homodimer in solution. On the other hand, values of the first-order rate constant for cyanide and imidazole dissociation from the diligated and monoligated derivatives of ferric Vitreoscilla Hb in solution are closely similar. As a whole, ligand binding and structural properties of ferric Vitreoscilla Hb appear to be unique among all Hbs investigated to date.

Selected figure(s)

Figure 3.
Figure 3. Kinetics for (a), (b) cyanide and (c), (d) imidazole binding to (a), (c) unligated and (b), (d) monoligated ferric Vitreoscilla Hb in solution at pH 7.0 and 20.0 °C. The cyanide and imidazole concentration refers to that of the free ligand. Continuous lines, representing the least-squares fitting of data, were obtained according to (a), (c) equation (2) and (b), (d) equation (3) with the following parameters. Cyanide: k[on] = 1.2(±0.2) × 10^2 M^−1 s^−1, α[on] = 3.3(±0.3) × 10^−3, k[off] = 2.3(±2.0) × 10^−4 s^−1 and α[off] = 1.0(±0.1). Imidazole: k[on] = 2.3(±0.3) × 10^4 M^−1 s^−1, α[on] = 3.3(±0.3) × 10^−3, k[off] = 9.3(±0.7) s^−1 and α[off] = 1.0(±0.1). For further experimental details, see the text.

Figure 5.
Figure 5. (a) Ribbon view of the Vitreoscilla Hb homodimer observed in the crystallographic asymmetric unit. In this (nearly symmetric) assembly the heme iron atoms are located about 34 Å apart. (b) Ribbon view of the crystal packing contacts affecting the heme distal and proximal regions of the A subunit cyan, taken as reference molecule from the asymmetric unit of Vitreoscilla Hb homodimer. The contact shown occurs with an equivalent molecule from the crystalline lattice (yellow), related to the subunit by a screw axis and by translations along a and c unit cell edges.

The above figures are reprinted by permission from Elsevier: J Mol Biol (1999, 291, 637-650) copyright 1999.

Figures were selected by an automated process.