H-bonding networks of the distal residues and water molecules in the active site of Thermobifida fusca hemoglobin

Interplay of the H-Bond Donor–Acceptor Role of the Distal Residues in Hydroxyl Ligand Stabilization of Thermobifida fusca Truncated Hemoglobin

“…When the OH group has strong hydrogen-bonding interactions with its neighboring groups, the expected isotope shift may be different because of the deviation from a perfect two-body 2 18 O, thus confirming the assignment to the ν(Fe− OH) stretching mode of the 6cLS form. 7 The corresponding band of the 6cHS has not been identified. The differences in the observed isotopic shifts with respect to the calculated values together with the remarkably low frequency, about 65 cm −1 lower than that observed for metMb and metHb, 8 indicates strong multiple hydrogen bonds between the OH − and distal residues.…”

“…Classical molecular dynamics simulations performed on the ASV ferric form reveal that the coordinated water is stabilized by strong H-bonds with the indole N proton of the WG8 (H 2 O−N WG8 = 1.9 Å) and the hydroxylic hydrogen of the YCD1 (O H 2 O −O YCD1 = 1.5 Å), confirming the spectroscopic findings that suggest a Fe−OH − character of the ligand even at pH 6.1. 7 The pK a of the acid−alkaline transition is an optimum probe of the overall charge distribution in the distal pocket microenvironment of heme proteins characterized by an aquo-met state at neutral pH. 8,12,13 In the case of Tf-trHb and its distal pocket mutants, the pK a of the acid−alkaline transition has been determined by absorption spectrophotometry.…”

“…Experimental conditions are as reported previously. 7 Absorption spectra were measured both prior to and after RR measurements at room temperature to ensure that no degradation had taken place under the experimental conditions used. All RR measurements were repeated several times under the same conditions to ensure reproducibility.…”

“…7 We found that the hydroxyl ligand does not bind to the triple mutant. In the case of the WT protein, binding gives rise to a sixcoordinate low-spin (6cLS) species with a ν(Fe−OH) stretching mode at 485 cm −1 , about 65 cm −1 lower than the corresponding bands observed for metMb and metHb 8−10 or any other truncated hemoglobin.…”

A Globin Domain in a Neuronal Transmembrane Receptor of Caenorhabditis elegans and Ascaris suum

“…A recent study on Thermobifida fusca hemoglobin revealed the presence of two ferric forms with similar EPR parameters as those found here. The component with a g 1 value around 2.8 was assigned to the hydroxide-ligated ferric complex where the OH group is strongly hydrogen bonded, whereas the form characterized by g 1 at ϳ2.6 is ascribed to a coordination mode in which the OH group is not so strongly H-bonded (79). Interestingly, also in T. fusca hemoglobin, the hydroxide-ligated complexes were still observable at pH 6.0.…”