Threonine Phosphorylation of the MMAC1/PTEN PDZ Binding Domain Both Inhibits and Stimulates PDZ Binding
2000
Two-hybrid searches with the tumor suppressor MMAC1/PTEN isolated the
proteins hDLG and hMAST205. Further two-hybrid analysis and microtiter
plate binding assays localized the sites of interaction to PDZ domains
from hDLG and hMAST205 and the PDZ binding domain at the COOH terminus
of MMAC1/PTEN. A synthetic peptide derived from the MMAC1/PTEN PDZ
binding domain (MMAC1/PTEN-PDZBD) was used to coprecipitate proteins
from A431 human cell lysate. The recovered proteins were resolved by
SDS-PAGE and immobilized on a nitrocellulose membrane. Treatment of
this membrane with an anti-hDLG antibody identified a
M r 140,000 band, consistent with the
size of hDLG. Treatment of this membrane with the MMAC1/PTEN-PDZBD
peptide identified a single prominent band of slightly larger than
M r 200,000 ( M r
200,000 kDa). Threonine phosphorylation of the MMAC1/PTEN-PDZBD peptide
inhibited both microtiter plate binding to the hDLG and hMAST205 PDZ
domains and coprecipitation of the M r
140,000 and >200,000 proteins, but promoted coprecipitation of
proteins of approximately M r 90,000 and
M r 120,000 from A431 cell lysate. This
result suggests phosphorylation of the MMAC1/PTEN PDZ binding domain
can both inhibit and promote PDZ interactions.
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