[Recombinant fragment of pigment epithelium-derived factor (44-77) prevents pathological corneal neovascularization].
Pigment epithelium-derived factor (PEDF), a 50 kDa secreted glycoprotein, is among the most potent endogenous inhibitors of angiogenesis. PEDF-derived fragment (44-77) possesses antiangiogenic properties of the full-sized protein and is a potential drug candidate for the treatment of ocular neovascular diseases. In this study we propose an efficient scalable biotechnological method for the production of PEDF (44-77) as part of a fusion protein with SspDnaB intein. The fusion protein was obtained in bacterial E. coli cells in the form of inclusion bodies, solubilized and subjected to autocatalytic cleavage with the release of PEDF (44-77) (yield, 77%). The target peptide was separated from the intein using tangential ultrafiltration. The final purification of PEDF (44-77) was performed by reversed-phase HPLC. The yield of the target peptide (purity, 99%) was 65 mg per 1 liter of culture. Antiangiogenic activity of the obtained peptide was studied in vitro using murine endothelial cells SVEC-4-10. PEDF (44-77) suppressed proliferation of endothelial cells by 53% and inhibited endothelial cell tube formation at the concentration of 1 nM. The ability of the recombinant PEDF (44-77) to block initial stages of angiogenesis was demonstrated using the model of rabbit corneal neovascularization.
R S
RS
Esipov
K A
KA
Beĭrakhova
L A
LA
Chupova
V K
VK
Likhvantseva
E V
EV
Stepanova
A I
AI
Miroshnikov
D020533:Angiogenesis Inhibitors / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D000818:Animals; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D016510:Corneal Neovascularization / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000378:metabolism / Q000473:pathology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005136:Eye Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D006801:Humans; D051379:Mice; D009414:Nerve Growth Factors / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology; D015843:Serpins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology*; D000099174:Pigment Epithelium-Derived Factor
D020533:Angiogenesis Inhibitors; D005136:Eye Proteins; D009414:Nerve Growth Factors; D011994:Recombinant Proteins; D015843:Serpins; D000099174:Pigment Epithelium-Derived Factor