tailieunhanh - Báo cáo khoa học: Pigment epithelium-derived factor binds to cell-surface F1-ATP synthase

Pigment epithelium-derived factor (PEDF), a potent blocker of angiogene-sis in vivo, and of endothelial cell migration and tubule formation, binds with high affinity to an as yet unknown protein on the surfaces of endothe-lial cells. Given that protein fingerprinting suggested a match of a 60 kDa PEDF-binding protein in bovine retina withBos taurusF1-ATP synthase b-subunit, and that F1Fo-ATP synthase components have been identified recently as cell-surface receptors, we examined the direct binding of PEDF to F1. . | Pigment epithelium-derived factor binds to cell-surface F1-ATP synthase I 111 Pi I M oÝĩỉ ri1 M I Aralpalpi1 2 11 o 11 o r3 Qmtt l l dio r3 111 a n A m a ra 11 Q 1 d R p Racnrra1 LUigi NUiali NaUkalU AlakaM David iviueilcl OCUll IVIclel JUaU Alllalal aMU S. I. oecella 1 Section of Protein Structure and Function Laboratory of RetinalCelland Molecular Biology NationalEye Institute NIH Bethesda MD USA 2 The University of Tokushima Graduate School Japan 3 Department of Biochemistry and Molecular Biology Rosalind Franklin University of Medicine and Science The Chicago Medical School IL USA Keywords endothelialcells F1-ATPase F1Fo-ATP synthase PEDF surface plasmon resonance Correspondence S. P. Becerra NIH-NEI Building 6 Room 134 6 Center Drive Bethesda MD 208920608 USA Fax 1 301 451 5420 Tel 1 301 496 6514 E-mail becerrap@ Website http intramural protein_struct_func Received 5 October 2009 revised 25 January 2010 accepted 3 March 2010 doi Pigment epithelium-derived factor PEDF a potent blocker of angiogenesis in vivo and of endothelial cell migration and tubule formation binds with high affinity to an as yet unknown protein on the surfaces of endothelial cells. Given that protein fingerprinting suggested a match of a 60 kDa PEDF-binding protein in bovine retina with Bos taunts F1-ATP synthase b-subunit and that F1Fo-ATP synthase components have been identified recently as cell-surface receptors we examined the direct binding of PEDF to F1. Size-exclusion ultrafiltration assays showed that recombinant human PEDF formed a complex with recombinant yeast F1. Realtime binding as determined by surface plasmon resonance demonstrated that yeast F1 interacted specifically and reversibly with human PEDF. Kinetic evaluations revealed high binding affinity for PEDF in agreement with PEDF affinities for endothelial cell surfaces. PEDF blocked interactions between F1 and angiostatin another antiangiogenic factor suggesting .