THE PL(A) ALLOANTIGEN SYSTEM IS A SENSITIVE INDICATOR OF THE STRUCTURAL INTEGRITY OF THE AMINO-TERMINAL DOMAIN OF THE HUMAN INTEGRIN BETA(3) SUBUNIT

Within the native integrin alpha(IIb)beta(3), the conformation of the amino-terminal domain of the beta(3) subunit has a significant influen ce on the availability of the Leu(33)/Pro(33) polymorphism that define s the Al and A2 alleles, respectively, of the Pl(A) alloantigen system . The majority of anti-Pl(Al) IgG antibodies, affinity-purified by ads orption to either (Al/Al)-platelets or purified alpha(IIb)(Al)beta(3), fail to bind to the Leu(33) polymorphic loop within Cys(26)-Cys(38) i n native beta(3) unless this sequence is maintained in a proper orient ation by two noncontiguous human pg sequences. By comparing IgG bindin g to recombinant beta(3) molecules composed of segments of human, Xeno pus and avian sequences expressed by Spodoptera frugiperda cell lines, we have found that neither region alone is sufficient for full expres sion of the epitope. One sequence, that most proximal to Leu(33)/Pro(3 3), lies within beta(3)[54-133], and a second distal sequence is locat ed within beta(3)[435-490]. Proximity of the distal sequence to the Cy s(26)-Cys(38) loop in native beta(3) is confirmed by the fact that bot h anti-Pl(Al) IgG and the murine monoclonal antibody SZ21, specific fo r beta(3)[28-35], completely inhibit the binding of rabbit polyclonal IgG specific for the sequence beta(3) [479-485]. Anti-Pl(Al) IgG antib odies from all of 7 donors selectively bind to the epitope within the native conformation of beta(3). However, in one of the seven donors, r oughly 60% of the anti-Pl(Al)IgG binds exclusively to denatured (Al)be ta(3), and not to either denatured (A2)beta(3) or nondenatured (native ) (Al)beta(3). This IgG subpopulation cannot be purified by adsorption to intact (Al/Al)-platelets or to native alpha(IIb)(Al)beta(3) integr in. The physiologic relevance of this exceptional type of anti-Pl(Al) antibody is debatable, but the presence of IgG with this specificity o nly in an immunized subject suggests that the epitope may well be pres ented on a ''denatured'' form of beta(3) in vivo. Regardless, our resu lts indicate that the amino-terminal domain of the native beta(3) mole cule associates with noncontiguous beta(3) sequences, and the epitope recognized by a majority of anti-Pl(Al) antibodies is a sensitive indi cator of correct tertiary structure within this region of beta(3).