Characterization of the WW domain of human yes-associated protein and its polyproline-containing ligands.
We had previously identified the WW domain as a novel globular domain that is composed of 38-40 semiconserved amino acids and is involved in mediating protein-protein interaction. The WW domain is shared by proteins of diverse functions including structural, regulatory, and signaling proteins in yeast, nematode, and mammals. Functionally it is similar to the Src homology 3 domain in that it binds polyproline ligands. By screening a 16-day mouse embryo expression library, we identified two putative ligands of the WW domain of Yes kinase-associated protein which we named WW domain-binding proteins 1 and 2. These proteins interacted with the WW domain via a short proline-rich motif with the consensus sequence of four consecutive prolines followed by a tyrosine. Herein, we report the cDNA cloning and characterization of the human orthologs of WW domain-binding proteins 1 and 2. The products encoded by these cDNA clones represent novel proteins with no known function. Furthermore, these proteins show no homology to each other except for a proline-rich motif. By fluorescence in situ hybridization on human metaphase chromosomes, we mapped the human genes for WW domain-binding proteins 1 and 2 to chromosomes 2p12 and 17q25, respectively. In addition, using site-directed mutagenesis, we determined which residues in the WW domain of Yes kinase-associated protein are critical for binding. Finally, by synthesizing peptides in which the various positions of the four consecutive proline-tyrosine motif and the five surrounding residues were replaced by all possible amino acid residues, we further elucidated the binding requirements of this motif.
Chen
H I
HI
Laboratory of Molecular Oncology, The Rockefeller University, New York, New York 10021, USA.
Einbond
A
A
Kwak
S J
SJ
Linn
H
H
Koepf
E
E
Peterson
S
S
Kelly
J W
JW
Sudol
M
M
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D018797:Cell Cycle Proteins; D002874:Chromosome Mapping; D002886:Chromosomes, Human, Pair 17; D002889:Chromosomes, Human, Pair 2; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D015723:Gene Library; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D047908:Intracellular Signaling Peptides and Proteins; D008024:Ligands; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010455:Peptides / Q000378:metabolism*; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D017433:Protein Structure, Secondary; D016415:Sequence Alignment; D015534:Trans-Activators; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008024:Ligands; D010455:Peptides; D010750:Phosphoproteins; D015534:Trans-Activators; D014157:Transcription Factors; C106907:WBP1 protein, human; C106908:WBP2 protein, human; C484574:Wbp1 protein, mouse; C484576:Wbp2 protein, mouse; D000091102:YAP-Signaling Proteins; C088374:YAP1 protein, human; C486223:Yap1 protein, mouse; C011083:polyproline