Random peptide bacteriophage display as a probe for urokinase receptor ligands.
The urokinase receptor is a multi-functional protein that plays a central role in cell surface plasminogen activation, cell migration, and cell adhesion. We previously demonstrated that high affinity peptide ligands for the urokinase receptor, which are urokinase competitors, can be obtained from a 15mer peptide library (Goodson et al., 1994). In order to probe for additional urokinase receptor binding sites we affinity selected the same bacteriophage library on complexes of soluble urokinase receptor (suPAR) and the receptor binding domain of urokinase, residues 1-48 (uPA1-48). Bacteriophage were isolated which bound to suPAR and suPAR:uPA1-48 complexes with high yield. The peptide sequences encoded by these bacteriophage were distinct from those obtained previously on urokinase receptor expressing cells, and comprise two groups based upon effects on su-PAR:1-anilino-8-napthalene sulfonate (ANS) fluorescence, and vitronectin binding competition. Alanine scanning mutagensis of the soluble peptides was used to define minimal regions and key residues for suPAR binding by competition with the parent bacteriophage. A comparison of these results with sequences of domains of both vitronectin and integrin alpha-chains, which have been reported to be important for urokinase receptor binding, suggests that the homology with the peptide sequences selected is functionally significant.
Fong
Susan
S
Chiron Corporation, Emeryville, CA 94608, USA.
Doyle
Michael V
MV
Goodson
Robert J
RJ
Drummond
Robert J
RJ
Stratton
Jennifer R
JR
McGuire
Lisa
L
Doyle
Laura V
LV
Chapman
Harold A
HA
Rosenberg
Steven
S
D000409:Alanine; D000595:Amino Acid Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D006801:Humans; D016023:Integrins / Q000378:metabolism; D008024:Ligands; D016296:Mutagenesis; D009842:Oligopeptides / Q000737:chemistry* / Q000378:metabolism; D019151:Peptide Library; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000378:metabolism; D055293:Receptors, Urokinase Plasminogen Activator; D012995:Solubility; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism; D019096:Vitronectin / Q000378:metabolism
D016023:Integrins; D008024:Ligands; D009842:Oligopeptides; C524898:PLAUR protein, human; D019151:Peptide Library; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D019096:Vitronectin; D014568:Urokinase-Type Plasminogen Activator; D000409:Alanine