Protonation states of histidine and other key residues in deoxy normal human adult hemoglobin by neutron protein crystallography

• Published in: Acta crystallographica. Section D, Biological crystallography. Vol. 66; no. 11; pp. 1144 - 1152
• Main Authors: Kovalevsky, Andrey, Chatake, Toshiyuki, Shibayama, Naoya, Park, Sam-Yong, Ishikawa, Takuya, Mustyakimov, Marat, Fisher, S. Zoe, Langan, Paul, Morimoto, Yukio
• Format: Journal Article
• Language: English
• Published: 5 Abbey Square, Chester, Cheshire CH1 2HU, England International Union of Crystallography 01.11.2010
• Subjects: Adult; AFFINITY; Bohr effect; CHAINS; CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CRYSTALLOGRAPHY; deoxy form; deuterated water; Hemoglobin; Hemoglobins - chemistry; Hemoglobins - metabolism; HISTIDINE; Histidine - chemistry; Histidine - metabolism; Human; human hemoglobin; Humans; INTERACTIONS; INTERFACES; Models, Molecular; NEUTRON DIFFRACTION; neutron protein crystallography; NEUTRONS; OXYGEN; Oxygen - chemistry; Protein Conformation; Protein Subunits; Proteins; Protonation; protonation states; PROTONS; Research Papers; Residues; STABILITY
• ISSN: 1399-0047; 0907-4449; 1399-0047
• DOI: 10.1107/S0907444910025448

The protonation states of the histidine residues key to the function of deoxy (T‐state) human hemoglobin have been investigated using neutron protein crystallography. These residues can reversibly bind protons, thereby regulating the oxygen affinity of hemoglobin. By examining the OMIT Fo − Fc and 2Fo−Fc neutron scattering maps, the protonation states of 35 of the 38 His residues were directly determined. The remaining three residues were found to be disordered. Surprisingly, seven pairs of His residues from equivalent α or β chains, αHis20, αHis50, αHis58, αHis89, βHis63, βHis143 and βHis146, have different protonation states. The protonation of distal His residues in the α1β1 heterodimer and the protonation of αHis103 in both subunits demonstrates that these residues may participate in buffering hydrogen ions and may influence the oxygen binding. The observed protonation states of His residues are compared with their ΔpKa between the deoxy and oxy states. Examination of inter‐subunit interfaces provided evidence for interactions that are essential for the stability of the deoxy tertiary structure.