Functional properties of Hb Pasadena, alpha 2 beta 2 75(E 19) Leu replaced by Arg

Functional properties of Hb Pasadena (beta 75[E19]Leu replaced by Arg) as well as correlations of its structure-function relationships with those of normal hemoglobin are presented. The homeotropic and heterotropic properties of the isolated variant were investigated by automatic recording oxygen equilibrium analysis. These studies indicate an increase of 02 affinity but normal cooperativity. The decrease of 02 affinity in response to organic phosphate (DPG and IHP) was found to be normal. However, the cooperativity of Hb Pasadena is less affected by IHP than is that of Hb A. The alkaline Bohr effect was found to be somewhat decreased, although the temperature effect is nearly normal. The autoxidation rate is increased by a factor of nearly 2 compared to normal. These observations are consistent with the conclusion that the beta (E19)75 leucyl hydrophobic residue of Hb A associates with beta (A8)11 Val, beta (A12)15 Trp and beta (H11)133 Val to form a hydrophobic cluster between the A, E and H helics which thereby stabilizes the heme of the beta chain in the deoxy ferrous state. The experimental results also suggest that the amino acid substitution in Hb Pasadena mainly disrupts the interior conformation of the beta subunit without affecting subunit interfaces. Thus it is predicted that Hb Pasadena should have a normal crystal structure in either the liganded (R) or unliganded (T) state. Its increased 02 affinity can be explained partially by a decrease of proton binding and partially by a shift of the R:T equilibrium towards the R conformation.