Effects of intra- and intersubunit hydrogen bonds on the R-T transition in human hemoglobin as studied with alpha 42(C7) and beta 145(HC2) mutations.
To clarify the effects of specific inter- and intrasubunit hydrogen bonds on the R-T transition in human hemoglobin (Hb A), the recombination reaction of carbon monoxide with artificial mutant Hbs was measured and analyzed. One of the hydrogen bonds we focused on is formed between Tyr-42 alpha and Asp-99 beta in the alpha 1-beta 2 interface of Hb A, which is one of the hydrogen bonds characteristic of the T state. Hb His-42 alpha, in which Tyr-42 alpha is replaced by His to perturb this hydrogen bond, showed that the ligand-free R to T transition rate was decreased by 20-fold compared with that for Hb A. This mutation caused the destabilization of the transition state in the R to T quaternary structure change by about 7 kJ mol-1, indicating that the hydrogen bond between Tyr-42 alpha and Asp-99 beta plays a definite role in the R-T transition as well as in stabilization of the equilibrium T state. Hb Phe-145 beta, in which Tyr-145 beta is replaced by Phe and the intrasubunit hydrogen bond between Tyr-145 beta and Val-98 beta is lacking, also showed a slow R-T transition rate as observed in Hb His-42 alpha. The published crystallographic data suggest that this intrasubunit hydrogen bond stabilizes the transition state by reducing the freedom of motion of the C-terminus of the beta subunit and, thereby, facilitates the R-T transition.
Division of Molecular Engineering, Graduate School of Engineering, Kyoto University, Japan.
A
A
Togi
K
K
Ishimori
M
M
Unno
T
T
Konno
I
I
Morishima
G
G
Miyazaki
K
K
Imai
D000595:Amino Acid Sequence; D001224:Aspartic Acid; D002248:Carbon Monoxide / Q000378:metabolism; D004926:Escherichia coli; D006441:Hemoglobin A / Q000737:chemistry* / Q000378:metabolism; D006639:Histidine; D006801:Humans; D006860:Hydrogen Bonding; D007700:Kinetics; D046911:Macromolecular Substances; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011487:Protein Conformation; D013816:Thermodynamics; D014443:Tyrosine
D046911:Macromolecular Substances; D001224:Aspartic Acid; D014443:Tyrosine; D006639:Histidine; D002248:Carbon Monoxide; D006441:Hemoglobin A