Human hemoglobin (HbA) is an intricate system that has evolved to efficiently transport oxygen molecules (O(2)) from lung to tissue. Its quaternary structure can fluctuate between two conformations, T (tense or deoxy) and R (relaxed or oxy), which have low and high affinity for O(2), respectively. The binding of O(2) to the heme sites of HbA is regulated by protons and by inorganic anions. In order to investigate the role of the protonation states of protein residues in O(2) binding, large crystals of deoxy HbA (approximately 20 mm(3)) were grown in D(2)O under anaerobic conditions for neutron diffraction studies. A time-of-flight neutron data set was collected to 1.8 A resolution on the Protein Crystallography Station (PCS) at the spallation source run by Los Alamos Neutron Science Center (LANSCE). The HbA tetramer (64.6 kDa; 574 residues excluding the four heme groups) occupies the largest asymmetric unit (space group P2(1)) from which a high-resolution neutron data set has been collected to date.