Visualizing the Bohr effect in hemoglobin: neutron structure of equine cyanomethemoglobin in the R state and comparison with human deoxyhemoglobin in the T state

Dajnowicz, Steven; Seaver, Sean; Hanson, B. Leif; Fisher, S. Zoë; Langan, Paul; Kovalevsky, Andrey Y.; Mueser, Timothy C.

doi:10.1107/s2059798316009049

Title: Visualizing the Bohr effect in hemoglobin: neutron structure of equine cyanomethemoglobin in the R state and comparison with human deoxyhemoglobin in the T state

Journal Article · Tue Jun 28 00:00:00 EDT 2016 · Acta Crystallographica. Section D. Structural Biology

DOI:https://doi.org/10.1107/s2059798316009049· OSTI ID:1625838

Dajnowicz, Steven ^[1]; Seaver, Sean ^[1]; Hanson, B. Leif ^[1]; Fisher, S. Zoë ^[2]; Langan, Paul ^[3]; Kovalevsky, Andrey Y. ^[3]; Mueser, Timothy C. ^[1]

Univ. of Toledo, OH (United States). Dept. of Chemistry and Biochemistry
European Spallation Source (ESS), Lund (Sweden). Scientific Activities Division. Science Directorate
Univ. of Toledo, OH (United States). Dept. of Chemistry and Biochemistry; Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biology and Soft Matter Division

Neutron crystallography provides direct visual evidence of the atomic positions of deuterium-exchanged H atoms, enabling the accurate determination of the protonation/deuteration state of hydrated biomolecules. Comparison of two neutron structures of hemoglobins, human deoxyhemoglobin (T state) and equine cyanomethemoglobin (R state), offers a direct observation of histidine residues that are likely to contribute to the Bohr effect. Previous studies have shown that the T-state N-terminal and C-terminal salt bridges appear to have a partial instead of a primary overall contribution. Four conserved histidine residues [αHis72(EF1), αHis103(G10), αHis89(FG1), αHis112(G19) and βHis97(FG4)] can become protonated/deuterated from the R to the T state, while two histidine residues [αHis20(B1) and βHis117(G19)] can lose a proton/deuteron. αHis103(G10), located in the α₁:β₁dimer interface, appears to be a Bohr group that undergoes structural changes: in the R state it is singly protonated/deuterated and hydrogen-bonded through a water network to βAsn108(G10) and in the T state it is doubly protonated/deuterated with the network uncoupled. The very long-term H/D exchange of the amide protons identifies regions that are accessible to exchange as well as regions that are impermeable to exchange. The liganded relaxed state (R state) has comparable levels of exchange (17.1% non-exchanged) compared with the deoxy tense state (T state; 11.8% non-exchanged). Interestingly, the regions of non-exchanged protons shift from the tetramer interfaces in the T-state interface (α₁:β₂and α₂:β₁) to the cores of the individual monomers and to the dimer interfaces (α₁:β₁and α₂:β₂) in the R state. The comparison of regions of stability in the two states allows a visualization of the conservation of fold energy necessary for ligand binding and release.

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Research Organization:: Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)

Sponsoring Organization:: USDOE Office of Science (SC)

Grant/Contract Number:: AC05-00OR22725

OSTI ID:: 1625838

Journal Information:: Acta Crystallographica. Section D. Structural Biology, Vol. 72, Issue 7; ISSN 2059-7983

Publisher:: IUCrCopyright Statement

Country of Publication:: United States

Language:: English

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59 BASIC BIOLOGICAL SCIENCES
Biochemistry & Molecular Biology
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neutron crystallography
cyanomethemoglobin
alkaline Bohr effect
H/D exchange