Stable interdomain interaction within the cytoplasmic domain of CD45 increases enzyme stability

CD45 is a leukocyte-specific, two domain transmembrane tyrosine phosphatase . Go-purification of a recombinant protein containing the first phosphatase domain of CD45 (6His-D1) with a recombinant protein containing the second phosphatase domain (GST-D2) from E. coli indicated a stable interaction whi ch resulted in increased stability of the active phosphatase domain present in 6His-D1. This interaction was not dependent on the acidic region unique to CD45 domain 2, but was affected by a destabilizing point mutation (Q118 0G) in GST-D2. CD45 domain 2 enhanced phosphatase activity of the first dom ain in the full length cytoplasmic domain protein, whereas a chimeric prote in with the SH2 domain of p56(lck) in place of the CD45 C-terminal region d id not. Thus the C-terminal domain of CD45 associates with the N-terminal d omain and this stabilizes the active phosphatase domain. A single destabili zing point mutation in the second domain is sufficient to attenuate this ef fect. (C) 2000 Academic Press.