Bioinformatic Characterization and Molecular Evolution of the Lucina pectinata Hemoglobins

Abstract

(1) Introduction: Lucina pectinata is a clam found in sulfide-rich mud environments that has three hemoglobins believed to be responsible for the transport of hydrogen sulfide (HbILp) and oxygen (HbIILp and HbIIILp) to chemoautotrophic endosymbionts. The physiological roles and evolution of these globins in sulfide-rich environments are not well understood. (2) Methods: We performed bioinformatic and phylogenetic analyses with 32 homologous mollusk globin sequences. Phylogenetics suggests a first gene duplication resulting in sulfide binding and oxygen binding genes. A more recent gene duplication gave rise to the two oxygen-binding hemoglobins. Multidimensional scaling analysis of the sequence space shows evolutionary drift of HbIILp and HbIIILp, while HbILp was closer to the Calyptogena hemoglobins. Further corroboration is seen by conservation in the coding region of hemoglobins from L. pectinata compared to those from Calyptogena. (3) Conclusions: Presence of glutamine in position E7 in organisms living in sulfide-rich environments can be considered an adaptation to prevent loss of protein function. In HbILp a substitution of phenylalanine in position B10 is accountable for its unique reactivity towards H2S. It appears that HbILp has been changing over time, apparently not subject to functional constraints of binding oxygen, and acquired a unique function for a specialized environment.

Hydrogen Sulfide Ligation in Hemoglobin I of Lucina pectinata─A QM/MM and Local Mode Study

“…NH c Bond. According to previous findings, 17,31,32 the proton transfer between H 2 S coordinated to the heme group and the distal residue (Gln64 and His64δ) stabilizes the protein active site. However, as already indicated by the optimized geometries discussed above, this proton transfer is accompanied by another proton transfer that occurs between the distal residue and COO − of one propionate heme group involving the NH c bond (see Figures 1c,d and S2−S4 in Supporting Information), which additionally stabilizes the protein active site.…”

“…We also investigated in our study the hydrogen acceptor O/NH a bonds, which has been suggested in the literature as a key player for the HbI H 2 S ligation ,, with an emphasis on their relation to the SH a bond. As revealed by the data in Table , the O and N hydrogen acceptor atoms of the distal residues form strong O/NH a bonds with BSO values in the range 0.796–0.829, with typical OH or NH bond lengths.…”

“…In contrast, in protein mutations in which no proton transfer is able to occur (Group 2), the SH a bond has a more covalent (the average H ρ = −0.200 hartree/ bohr 3 ) and stronger bond character, while the FeS bond is weakened and becomes significantly much less covalent (the average H ρ = −0.011 hartree/bohr 3 ). We also investigated in our study the hydrogen acceptor O/ NH a bonds, which has been suggested in the literature as a key player for the HbI H 2 S ligation 17,31,32 with an emphasis on their relation to the SH a bond. As revealed by the data in Table 4, the O and N hydrogen acceptor atoms of the distal residues form strong O/NH a bonds with BSO values in the range 0.796−0.829, with typical OH or NH bond lengths.…”

“…The properties of H 2 S coordinated to HbI and the role of the surrounding amino acids and their mutations involving distal glutamine and Phe forming the Phe cage around H 2 S in Lucina pectinata were investigated using experimental methods such as X-ray, resonance Raman measurements, UV–vis spectroscopy, and fluorescence spectroscopy . The kinetics and dynamics of HbI ligand binding were experimentally investigated ,, and complemented with theoretical studies utilizing hybrid molecular dynamics and combined quantum mechanical–molecular mechanical (QM/MM) calculations. ,,, Through these studies, they have found that the presence of distal Gln can be considered an adaptation to prevent loss of protein function for organisms living in a sulfide-rich environment, such as in the Lucina pectinata, and it has been argued that the distal Gln, along with the Phe cage, is critical for the stabilization and release of H 2 S through hydrogen bonding, i.e., the strength of the hydrogen bond between the distal residue and the bound ligand regulates H 2 S dissociation. ,, Whereas these studies have led to important overall insights, a detailed analysis of the mechanism at the molecular level including the assessment of the intrinsic strength of these hydrogen bonds has been missing so far, as well as that of the FeS bond, which is also a key player in this process.…”

“…17,21,24,31 Through these studies, they have found that the presence of distal Gln can be considered an adaptation to prevent loss of protein function for organisms living in a sulfide-rich environment, such as in the Lucina pectinata, and it has been argued that the distal Gln, along with the Phe cage, is critical for the stabilization and release of H 2 S through hydrogen bonding, i.e., the strength of the hydrogen bond between the distal residue and the bound ligand regulates H 2 S dissociation. 17,31,32 Whereas these studies have led to important overall insights, a detailed analysis of the mechanism at the molecular level including the assessment of the intrinsic strength of these hydrogen bonds has been missing so far, as well as that of the FeS bond, which is also a key player in this process.…”