Hemoglobin I (HbI) from the clam Lucina pectinata is, in its natural environment, a hydrogen sulfide (H2S)-transport heme protein. The resonance Raman (RR) spectrum of the metaquo and deoxyHbI species shows a very weak intensity peak at 370 cm-1 that corresponds to the normal mode of the heme propionates. This suggests the presence of a moderate hydrogen bonding between Arg99 and the heme-7-propionate. However, the RR spectra of the metcyano, carbonmonoxy, and oxy HbI derivatives reveal the absence of the propionate vibrational frequency at 370 cm-1. The mode is insensitive to the oxidation state of the heme iron, but disappears when the HbI−ligand moiety is formed. These results propose the existence of flexible propionate groups which can result in a weaker hydrogen bond upon heme ligand binding. The longitudinal relaxation time (T 1) 1H NMR data for the paramagnetic metcyano complex of HbI suggested that the 17.90 ppm signal belongs to the heme-6-propionate α‘ protons (6-Hα ‘). In relation to other myoglobins, the large difference in chemical shifts of this signal is attributed both to the lack of hydrogen bonds between the heme-6-propionate group and amino acid residues and to a flexible orientation of the side chain with respect to the heme plane. The data predict a model where the heme group of HbI is tightly bound to His96 (νFe - His at 218 cm-1), but due to the absence of strong hydrogen bonding interactions between the heme propionates and the nearby amino acids, the heme is not firmly anchored. Thus, relative to other heme proteins, the heme group of HbI from Lucina pectinata presents a rocking freedom that facilitates the binding between the heme and the in-coming ligand.

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