Characterization of the interaction between RhoA and the amino-terminal region of PKN.
The yeast two-hybrid system and in vitro binding assay were carried out to characterize the interaction between PKN and a small GTP-binding protein, RhoA. It was revealed that the region corresponding to the amino acid residues 33-111 in the amino-terminal region of PKN was sufficient to confer the ability to associate with RhoA. Each synthetic peptide fragment corresponding to the amino acid residues 74-93 and 94-113 of PKN inhibited the interaction between PKN and RhoA in the in vitro binding assay, suggesting that this region is important in the association with RhoA. The endogenous and the GAP-stimulated GTPase activity of RhoA was inhibited by the interaction with PKN, suggesting the presence of a regulatory mechanism that sustains the GTP-bound active form of RhoA.
Shibata
H
H
Department of Biology, Faculty of Science, Kobe University, Japan.
Mukai
H
H
Inagaki
Y
Y
Homma
Y
Y
Kimura
K
K
Kaibuchi
K
K
Narumiya
S
S
Ono
Y
Y
D001665:Binding Sites; D020558:GTP Phosphohydrolases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D005822:Genetic Vectors; D005982:Glutathione Transferase / Q000235:genetics; D006160:Guanosine Triphosphate / Q000378:metabolism; D016350:Leucine Zippers; D009154:Mutation; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D011506:Proteins / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D014162:Transfection; D020742:rhoA GTP-Binding Protein
D020690:GTPase-Activating Proteins; D010446:Peptide Fragments; D011506:Proteins; D011993:Recombinant Fusion Proteins; D006160:Guanosine Triphosphate; D005982:Glutathione Transferase; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein