The spectrin heterodimer is formed by the antiparallel lateral association of an 
alpha and a beta subunit, each of which comprises largely a series of homologous 
triple-helical motifs. Initiation of dimer assembly involves strong binding 
between complementary motifs near the actin-binding end of the dimer. In this 
study, the mechanism of lateral spectrin association at this dimer nucleation 
site was investigated using the analytical ultracentrifuge to analyze 
heterodimers formed from recombinant peptides containing two or four homologous 
motifs from each subunit (alpha20-21/beta1-2; alpha18-21/beta1-4). Both the 
two-motif and four-motif dimer associations were weakened substantially with 
increasing salt concentration, indicating that electrostatic interactions are 
important for the dimer initiation process. Modeling of the electrostatic 
potential on the surface of the alpha20 and beta2 motifs showed that the side of 
the motifs comprising the A and B helices is the most favorable for association, 
with an area of positive electrostatic potential on the AB face of the beta2 
motif opposite negative potential on the AB face of the alpha20 motif and vise 
versa. Protease protection analysis of the alpha20-21/beta1-2 dimer showed that 
multiple trypsin and proteinase K sites in the A helices of the beta2 and 
alpha21 motifs become buried upon dimer formation. Together, these data support 
a model where complementary long range electrostatic interactions on the AB 
faces of the triple-helical motifs in the dimer nucleation site initiate the 
correct pairing of motifs, i.e. alpha21-beta1 and alpha20-beta2. After initial 
docking of these complementary triple-helical motifs, this association is 
probably stabilized by subsequent formation of stronger hydrophobic interactions 
in a complex involving the A helices of both subunits and possibly most of the 
AB faces. The beta subunit A helix in particular appears to be buried in the 
dimer interface.