MODULE SUBSTITUTION IN HEMOGLOBIN SUBUNITS - PREPARATION AND CHARACTERIZATION OF A CHIMERA BETA-ALPHA-SUBUNIT

In the genes of alpha- and beta-subunits of hemoglobin, Go showed that modules F1, F2 + F3, and F4 correspond to exons 1, 2, and 3, respecti vely (Go, M. (1981) Nature 291, 90). The analysis of the correlation o f function with its exon pattern showed that the residues associated w ith the defined function are concentrated in the specific exons encodi ng the ''module'' (Eaten, W.A. (1980) Nature 284, 183). To investigate the functional and structural significance of the ''modular structure ,'' we engineered a ''chimera'' subunit, in which module F4 of the bet a-subunit was replaced by that of the alpha-subunit by use of mutagene sis. The NMR and resonance Raman spectra of the isolated ''chimera bet a alpha-subunit'' have revealed that it has a beta-subunit-like heme e nvironmental structure. However, the gel chromatography and NMR spectr a of mixtures of the chimera and native subunits clearly showed that t he chimera beta alpha-subunit binds specifically to the beta-subunit t o form a heterotetramer, not to the alpha-subunit. These results led u s to conclude that the predominant role of the module F4 is the subuni t association and suggest that the modules are structural and function al units that have advantages in producing stable functional proteins.