Transposing sequences between fetal and adult hemoglobins indicates which subunits and regulatory molecule interfaces are functionally related.
To correlate amino acid sequence changes with hemoglobin function we are carrying out a detailed recombinant analysis of the adult hemoglobin/fetal hemoglobin (HbA/HbF) systems. The important physiological differences between these two tetramers lie at unspecified sites in the 39 sequence substitutions of the 146 amino acids in their beta and gamma chains. In this paper, significant differences in the tetramer-dimer dissociation constants (referred to as tetramer "strength" or "stability") of adult (HbA) and fetal (HbF) hemoglobin tetramers have been used to probe the relationship between the allosteric, sliding interface and the effects of the allosteric regulator, 2,3-DPG, in promoting oxygen release. The single amino acid difference at the allosteric interfaces of these two hemoglobins, Glu-43(beta) --> Asp-43(gamma), which is not near the DPG binding site, leads to a significantly lower DPG response, approaching that of HbF. The results are inconsistent with the long-held idea that the replacement of His-143(beta) in HbA to Ser-143(gamma) in HbF is solely responsible for the lowered DPG response in HbF. On the other hand, the Val-1(beta) --> Gly-1(gamma) replacement near the DPG binding site has no effect on the DPG response. The replacement of His-116(beta) by the hydrophobic Ile-116(gamma) at the rigid alpha(1)beta(1) interface has a marginal yet detectable effect on the allosteric alpha(1)beta(2) interface. The results, overall, are interpreted using a model involving electrostatic coupling between certain side chains and extend the concept of a long-range relationship between some distant regions of the tetramer that are likely mediated through the central cavity.
Chen
W
W
Department of Biology, Northeastern University, Boston, MA 02115, USA.
Dumoulin
A
A
Li
X
X
Padovan
J C
JC
Chait
B T
BT
Buonopane
R
R
Platt
O S
OS
Manning
L R
LR
Manning
J M
JM
D019794:2,3-Diphosphoglycerate / Q000378:metabolism; D000328:Adult; D000494:Allosteric Regulation / Q000235:genetics; D000595:Amino Acid Sequence; D001665:Binding Sites / Q000235:genetics; D019281:Dimerization; D005319:Fetal Hemoglobin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D005998:Glycine / Q000235:genetics; D006441:Hemoglobin A / Q000235:genetics; D006454:Hemoglobins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010100:Oxygen / Q000378:metabolism; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014633:Valine / Q000235:genetics
D006454:Hemoglobins; D011994:Recombinant Proteins; D019794:2,3-Diphosphoglycerate; D006441:Hemoglobin A; D005319:Fetal Hemoglobin; D014633:Valine; D010100:Oxygen; D005998:Glycine