Ligand Migration and Cavities within Scapharca Dimeric HbI: Studies by Time-Resolved Crystallo- graphy, Xe Binding, and Computational Analysis

Knapp, James E.; Pahl, R.; Cohen, Joseph R.; Nichols, Jeffry C.; Schulten, Klaus; Gibson, Quentin H.; Šrajer, V.; Royer, William E.

doi:10.1016/j.str.2009.09.004

Cited by 60 publications

(85 citation statements)

References 45 publications

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“…The second Leu16 conformation is associated with a smaller rotation of the 7-propionate. Significantly, this conformer exhibits a significantly bent Fe-C-O unit with a bond angle of 158 AE 8°( Table 1) comparable to that found in a variety of haemoproteins at atomic resolution structures (7)(8)(9)(10)(11)(12).…”

Section: Resultssupporting

confidence: 69%

Carbon monoxide poisoning is prevented by the energy costs of conformational changes in gas-binding haemproteins

Antonyuk

Rustage

Petersen

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Carbon monoxide (CO) is a product of haem metabolism and organisms must evolve strategies to prevent endogenous CO poisoning of haemoproteins. We show that energy costs associated with conformational changes play a key role in preventing irreversible CO binding. AxCYTcp is a member of a family of haem proteins that form stable 5c-NO and 6c-CO complexes but do not form O 2 complexes. Structure of the AxCYTcp-CO complex at 1.25 Å resolution shows that CO binds in two conformations moderated by the extent of displacement of the distal residue Leu16 toward the haem 7-propionate. The presence of two CO conformations is confirmed by cryogenic resonance Raman data. The preferred linear Fe-C-O arrangement (170 AE 8°) is accompanied by a flip of the propionate from the distal to proximal face of the haem. In the second conformation, the Fe-C-O unit is bent (158 AE 8°) with no flip of propionate. The energetic cost of the CO-induced Leupropionate movements is reflected in a 600 mV (57.9 kJmol −1 ) decrease in haem potential, a value in good agreement with density functional theory calculations. Substitution of Leu by Ala or Gly (structures determined at 1.03 and 1.04 Å resolutions) resulted in a haem site that binds CO in the linear mode only and where no significant change in redox potential is observed. Remarkably, these variants were isolated as ferrous 6c-CO complexes, attributable to the observed eight orders of magnitude increase in affinity for CO, including an approximately 10,000-fold decrease in the rate of dissociation. These new findings have wide implications for preventing CO poisoning of gas-binding haem proteins.sensors | crystallography | kinetics | ligand discrimination | atomic resolution

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Section: Resultssupporting

confidence: 69%

Carbon monoxide poisoning is prevented by the energy costs of conformational changes in gas-binding haemproteins

Antonyuk

Rustage

Petersen

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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“…Even though xenon binding to animal Hbs and Mbs is well documented, the results for CerHb are the first to show a large effect on the fraction of geminate recombination and that the xenon cavities are on the pathway for ligand entry and escape. In contrast, xenon binding to sperm whale Mb and S. inaequivalvis Hb causes little or no change in F gem for either O 2 or CO rebinding (13,18,19).…”

Section: Discussionmentioning

confidence: 87%

“…However, in most cases there has been little experimental proof that these tunnels are used by diatomic ligands. Cavities large enough to accommodate xenon atoms have been found in most globins, including sperm whale Mb and Scapharca inaequivalvis HbI; however, in both of the latter cases, xenon binding has little effect on either the fraction of geminate recombination or on overall rates of ligand binding or release (13,18,19), implying that these internal spaces are not part of the route for ligand entry and escape.…”

mentioning

confidence: 99%

Ligand Migration in the Apolar Tunnel of Cerebratulus lacteus Mini-Hemoglobin

Pesce

Nardini

Dewilde

et al. 2011

Journal of Biological Chemistry

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The large apolar tunnel traversing the mini-hemoglobin from Cerebratulus lacteus (CerHb) has been examined by xray crystallography, ligand binding kinetics, and molecular dynamic simulations. The addition of 10 atm of xenon causes loss of diffraction in wild-type (wt) CerHbO 2 crystals, but Leu-86(G12)Ala CerHbO 2 , which has an increased tunnel volume, stably accommodates two discrete xenon atoms: one adjacent to Leu-86(G12) and another near Ala-55(E18). Molecular dynamics simulations of ligand migration in wt CerHb show a low energy pathway through the apolar tunnel when Leu or Ala, but not Phe or Trp, is present at the 86(G12) position. The addition of 10 -15 atm of xenon to solutions of wt CerHbCO and L86A CerHbCO causes 2-3-fold increases in the fraction of geminate ligand recombination, indicating that the bound xenon blocks CO escape. This idea was confirmed by L86F and L86W mutations, which cause even larger increases in the fraction of geminate CO rebinding, 2-5-fold decreases in the bimolecular rate constants for ligand entry, and large increases in the computed energy barriers for ligand movement through the apolar tunnel. Both the addition of xenon to the L86A mutant and oxidation of wt CerHb heme iron cause the appearance of an out Gln-44(E7) conformer, in which the amide side chain points out toward the solvent and appears to lower the barrier for ligand escape through the E7 gate. However, the observed kinetics suggest little entry and escape (<25%) through the E7 pathway, presumably because the in Gln-44(E7) conformer is thermodynamically favored. Although molecular dynamics (MD)7 simulations suggest multiple interior pathways for O 2 entry into and exit from globins, most experiments with mammalian myoglobins (Mbs) and hemoglobins (Hbs) suggest a well defined single pathway involving a short channel between the heme propionates and the heme iron atom that is gated by the distal E7 histidine (1). To search for and define an interior ligand migration trajectory, we chose to use the neuronal mini-hemoglobin from Cerebratulus lacteus as a model globin system to examine its long apolar tunnel that leads from the distal portion of the heme pocket to an exit point between the C-terminal regions of the E and H helices of the tertiary fold, a pathway that is roughly 180°opposite the E7 channel and appears to be a major route for ligand entry (2). This model globin provides a well defined system to examine both experimentally and theoretically the effects of xenon binding, mutagenesis, and conformational heterogeneity on the competition between movement through the E7 gate versus an internal apolar pathway.Nerve tissue Hbs occur in both vertebrates and invertebrates (3). Among these, the nerve Hb from the nemertean worm C. lacteus (CerHb) is the smallest functional globin known, being composed of 109 amino acids instead of the ϳ140 -160 residues typical of most monomeric globins (4, 5). Analysis of the three-dimensional structure of CerHb has shown a markedly edited 3-over-3-globin fold with deletion of

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“…Xenon, which has often been used to probe dioxygen transient pathway, binds within this cavity. For example, in globin and in enzymes, xenon binding sites in cavities map small gaseous ligands transient pathway [38,[47][48][49][50][51][52][53][54]. In UOX, the internal cavity where xenon binds is very close to the active site and can have the same function as in globin for dioxygen.…”

Section: Discussionmentioning

confidence: 99%

Functional relevance of the internal hydrophobic cavity of urate oxidase

Colloc’h

Prangé

2014

FEBS Letters

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a b s t r a c tUrate oxidase from Aspergillus flavus is a 135 kDa homo-tetramer which has a hydrophobic cavity buried within each monomer and located close to its active site. Crystallographic studies under moderate gas pressure and high hydrostatic pressure have shown that both gas presence and high pressure would rigidify the cavity leading to an inhibition of the catalytic activity. Analysis of the cavity volume variations and functional modifications suggest that the flexibility of the cavity would be an essential parameter for the active site efficiency. This cavity would act as a connecting vessel to give flexibility to the neighboring active site, and its expansion under pure oxygen pressure reveals that it might serve as a transient reservoir on its pathway to the active site.

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Ligand Migration and Cavities within Scapharca Dimeric HbI: Studies by Time-Resolved Crystallo- graphy, Xe Binding, and Computational Analysis

Cited by 60 publications

References 45 publications

Carbon monoxide poisoning is prevented by the energy costs of conformational changes in gas-binding haemproteins

Carbon monoxide poisoning is prevented by the energy costs of conformational changes in gas-binding haemproteins

Ligand Migration in the Apolar Tunnel of Cerebratulus lacteus Mini-Hemoglobin

Functional relevance of the internal hydrophobic cavity of urate oxidase

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