The pigment epithelium-derived factor (PEDF), a secreted 50 kDa glycoprotein, is one of the most potent endogenous inhibitors of angiogenesis. The fragment 44-77 of PEDF possesses the antiangiogenic properties of the full-sized protein and is a potential drug candidate for the treatment of diseases of visual organs accompanied by pathological neovascularization. An effective biotechnological method for the large-scale production of the PEDF (44-77) fragment as part of the fusion protein with the SspDnaB intein has been developed. The hybrid protein was produced in Escherichia coli bacterial cells in the form of inclusion bodies, solubilized, and subjected to autocatalytic cleavage with the release of the PEDF (44-77) fragment (reaction yield 77%). The target peptide was separated from the intein by 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 l of culture. The antiangiogenic activity of the peptide was confirmed in vitro using mouse endothelial cells SVEC-4-10. It was found that the preparation at a concentration of 1 nM suppresses the proliferation of endothelial cells by 53% and inhibits the formation of tube-like structures by endothelial cells. The ability of the recombinant PEDF (44-77) to block the initial stages of angiogenesis was shown using an experimental model of rabbit corneal neovascularization. © Pleiades Publishing, Ltd. 2012.