Interaction of the p85 subunit of PI 3-kinase and its N-terminal SH2 domain with a PDGF receptor phosphorylation site: structural features and analysis of conformational changes.
Circular dichroism and fluorescence spectroscopy were used to investigate the structure of the p85 alpha subunit of the PI 3-kinase, a closely related p85 beta protein, and a recombinant SH2 domain-containing fragment of p85 alpha. Significant spectral changes, indicative of a conformational change, were observed on formation of a complex with a 17 residue peptide containing a phosphorylated tyrosine residue. The sequence of this peptide is identical to the sequence surrounding Tyr751 in the kinase-insert region of the platelet-derived growth factor beta-receptor (beta PDGFR). The rotational correlation times measured by fluorescence anisotropy decay indicated that phosphopeptide binding changed the shape of the SH2 domain-containing fragment. The CD and fluorescence spectroscopy data support the secondary structure prediction based on sequence analysis and provide evidence for flexible linker regions between the various domains of the p85 proteins. The significance of these results for SH2 domain-containing proteins is discussed.
Panayotou
G
G
Ludwig Institute for Cancer Research, London, UK.
Bax
B
B
Gout
I
I
Federwisch
M
M
Wroblowski
B
B
Dhand
R
R
Fry
M J
MJ
Blundell
T L
TL
Wollmer
A
A
Waterfield
M D
MD
D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D005454:Fluorescence Polarization; D007313:Insecta; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D010770:Phosphotransferases / Q000737:chemistry / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000378:metabolism*; D017433:Protein Structure, Secondary; D017479:Receptors, Platelet-Derived Growth Factor / Q000737:chemistry / Q000378:metabolism*
D010446:Peptide Fragments; D010982:Platelet-Derived Growth Factor; D010770:Phosphotransferases; D019869:Phosphatidylinositol 3-Kinases; D017479:Receptors, Platelet-Derived Growth Factor