One of the most common DNA lesions is the appearance of apurinic/apyrimidinic (AP-) sites. The main repair pathway for AP sites is initiated by apurinic/apyrimidinic endonuclease 1 (APE1). Upon hydrolysis of the phosphodiester bond by this enzyme, a one nucleotide gap flanked by 3'-hydroxyl and 5'-deoxyribose phosphate groups on the 5'-side of the AP site is formed. After hydrolysis of the AP site, APE1 remains associated with the product for some time. In the present work, the ability of APE1 to form a product of covalent attachment of APE1 to DNA containing a gap with a 5'-deoxyribose phosphate residue was demonstrated. In addition, it was found that while in a complex with the product of hydrolysis of the AP site, APE1 exhibits 5'-deoxyribose phosphate lyase activity, cleaving off the 5'-deoxyribose phosphate residue. The presence of lyase activity in APE1 may be important for the repair of AP sites if there is a deficiency of, or mutations in DNA polymerase β, the main enzyme that removes the 5'-deoxyribose phosphate group.