Sm. Bradykalnay et Nk. Tonks, IDENTIFICATION OF THE HEMOPHILIC BINDING-SITE OF THE RECEPTOR PROTEIN-TYROSINE-PHOSPHATASE PTP-MU, The Journal of biological chemistry, 269(45), 1994, pp. 28472-28477
The receptor-type protein tyrosine phosphatase PTP mu comprises an ext
racellular segment containing a MAM domain, an immunoglobulin domain a
nd four fibronectin type III repeats, a transmembrane segment, and two
intracellular PTP domains. We have previously shown that PTP mu binds
homophilically, i.e. PTP mu on the surface of one cell binds to PTP m
u on an apposing cell, and that the extracellular segment alone is suf
ficient for hemophilic binding. In this study we report that in MvLu c
ells PTP mu is proteolytically processed into two noncovalently associ
ated fragments, one comprising most of the extracellular segment (simi
lar to 100 kDa) and the other containing predominantly the transmembra
ne and intracellular portions (similar to 100 kDa). We have also ident
ified the hemophilic binding site within the extracellular segment. We
have generated, expressed, and purified various fragments of the extr
acellular segment of PTP mu and have used fluorescent beads (Covaspher
es) coated with these fragments in three binding assays: (i) measureme
nt of bead aggregation, (ii) binding of beads to surfaces of dishes co
ated with purified PTP mu or (iii) binding to MvLu cells. Only beads c
oated with recombinant fragments that contained the immunoglobulin dom
ain underwent aggregation or bound to surfaces displaying PTP mu, sugg
esting that neither the MAM domain nor the fibronectin type III repeat
s bound homophilically in these assays. The fragment containing the Ig
domain alone bound as well as any other Ig domain-containing fragment
, suggesting that the Ig domain is both necessary and sufficient for h
emophilic binding under these conditions.