The role of the distal histidine in myoglobin and haemoglobin

Abstract

The distal E7 histidine in vertebrate myoglobins and haemoglobins has been strongly conserved during evolution and is thought to be important in fine-tuning the ligand affinities of these proteins. A hydrogen bond between the N epsilon proton of the distal histidine and the second oxygen atom may stabilize O2 bound to the haem iron. The proximity of the imidazole side chain to the sixth coordination position, which is required for efficient hydrogen bonding, has been postulated to inhibit sterically the binding of CO and alkyl isocyanides. To test these ideas, engineered mutants of sperm whale myoglobin and the alpha- and beta-subunits of human haemoglobin were prepared in which E7 histidine was replaced by glycine. Removal of the distal imidazole in myoglobin and the alpha-subunits of intact, R-state haemoglobin caused significant changes in the affinity for oxygen, carbon monoxide and methyl isocyanide; in contrast, the His-E7 to Gly substitution produced little or no effect on the rates and extents of O2, CO and methyl isocyanide binding to beta-chains within R-state haemoglobin. In the beta-subunit the distal histidine seems to be less significant in regulating the binding of ligands to the haem iron in the high affinity quaternary conformation. Structural differences in the oxygen binding pockets shown by X-ray crystallographic studies account for the functional differences of these proteins.

Slow Ligand Binding Kinetics Dominate Ferrous Hexacoordinate Hemoglobin Reactivities and Reveal Differences between Plants and Other Species

“…All of the reactions measured so far have been CO binding to the ferrous form of each protein. To be sure that this behavior is not specific to this ligand, we used methyl isocyanide (MeICN) as ferrous ligand that, like CO, binds in the presence of sodium dithionite (25). These reactions were conducted with Ngb and Cgb because their time courses should be completely limited by k -H and, thus, independent of ligand concentration and the k′ L value specific to each ligand.…”

Application of molecular dynamics and free energy perturbation methods to metalloporphyrin‐ligand systems ii: Co and dioxygen binding to myoglobin

“…Recently, site-specific mutagenesis has allowed for the expression of Mb and Hb mutants in Escherichia coli (Springer & Sligar, 1987). This has allowed functional and structural evaluations of the residues His 64(E7) (Olson et al, 1988;Springer et al, 1989;Rohlfs et al, 1990;Ikeda-Saito et al, 1991;Petrich et al, 1991), Val 68(Ell) Smerdon et al, 1991), Arg 45 or Lys 45(CD3) (Lambright et al, 1989;Czrver et al, 1991), and Leu 29(B10) (Adachi et al, 1992;Carver et al, 1992;Gibson et al, 1992) in mammalian Mb. Similar studies with mutant Hbs have also been reported (Nagai et al, 1987;Mathews et al, 1989Mathews et al, , 1991Tame et al, 1991).…”

Modulation of Protein Function by Exogenous Ligands in Protein Cavities: CO Binding to a Myoglobin Cavity Mutant Containing Unnatural Proximal Ligands

“…There is now an extensive literature on synthetic biomimetic heme complexes and naturally occurring or engineered myoglobin mutants (Momenteau & Reed, 1994;Springer et al, 1994;Traylor & Traylor, 1982). Much of the effort on both model heme complexes and site-directed mutants of myoglobin and hemoglobin is directed toward understanding the interactions between diatomic ligands such as O 2 , CO, and NO and the environment of the distal heme pocket (Collman et al, 1979;Nagai et al, 1987;Olson et al, 1988;Lambright et al, 1989Lambright et al, , 1994Egeberg et al, 1990;Balasubramanian et al, 1993a).…”