Direct binding of recombinant plasminogen kringle 1-3 to angiogenin inhibits angiogenin-induced angiogenesis in the chick embryo CAM.
Angiogenin is one of the most potent angiogenesis-inducing proteins. Angiostatin is one of the most potent angiogenesis inhibitors, and it contains the first four kringle domains of plasminogen (K1-4). Recombinant human plasminogen kringle 1-3 (rK1-3) was expressed in Escherichia coli and purified to homogeneity. The binding of t-4-aminomethylcyclohexanecarboxylic acid with the purified kringle 1-3 was determined by changes in intrinsic fluorescence. rK1-3 exhibits comparable ligand-binding properties as native human plasminogen kringle 1-3. The purified rK1-3 inhibits neovascularization in the chick embryo chorioallantoic membrane (CAM) assay. Interaction of angiogenin with rK1-3 was examined by immunological binding assay and surface plasmon resonance kinetic analysis, and the equilibrium dissociation constants for the complex, Kd, are 0.89 and 0.18 microM, respectively. rK1-3 inhibits angiogenin-induced angiogenesis in the chick embryo CAM in a concentration-dependent manner. These results indicate that rK1-3 directly binds to angiogenin and thus rK1-3 inhibits the angiogenic activity of angiogenin.
Department of Biochemistry, Chungbuk National University, Cheongju 361-763, Republic of Korea.
Mi-Ran
MR
Youn
Mee-Hee
MH
Park
Chang-Ki
CK
Choi
Byung-Cheol
BC
Ahn
Hak Yong
HY
Kim
Sang Sun
SS
Kang
Yong-Kil
YK
Hong
Young Ae
YA
Joe
Jong-Soo
JS
Kim
Weon-Kyoo
WK
You
Hyo-Sil
HS
Lee
Soo-Il
SI
Chung
Soo-Ik
SI
Chang
D020533:Angiogenesis Inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002642:Chick Embryo; D049033:Chorioallantoic Membrane / Q000098:blood supply*; D004797:Enzyme-Linked Immunosorbent Assay; D008024:Ligands; D018919:Neovascularization, Physiologic; D010446:Peptide Fragments / Q000302:isolation & purification / Q000378:metabolism*; D010958:Plasminogen / Q000302:isolation & purification / Q000378:metabolism*; D012259:Ribonuclease, Pancreatic / Q000037:antagonists & inhibitors* / Q000378:metabolism; D020349:Surface Plasmon Resonance
D020533:Angiogenesis Inhibitors; D008024:Ligands; D010446:Peptide Fragments; C402400:recombinant kringle 1-3 (rKI-3) of human plasminogen; D010958:Plasminogen; C046366:angiogenin; D012259:Ribonuclease, Pancreatic