Boehm T, O'reilly M S, Keough K, Shiloach J, Shapiro R, Folkman J
The Children's Hospital, and Departments of Surgery and Cellular Biology, Harvard Medical School, 300 Longwood Avenue, Boston, Massachusetts, 02115, USA.
Biochem Biophys Res Commun. 1998 Nov 9;252(1):190-4. doi: 10.1006/bbrc.1998.9617.
Endostatin is a potent angiogenesis inhibitor in vitro and in vivo. We used the yeast Pichia pastoris to express and purify soluble endostatin. It was discovered that metal chelating agents can induce N-terminal degradation of endostatin. We theorized that a metal was removed from endostatin which changed the conformation and allowed a contaminating protease to degrade the N-terminus. Atomic absorption and amino acid analysis of endostatin purified from Pichia pastoris and mammalian cells showed a 1:1 molar ratio of Zn2+ to protein. Ding et al. have shown that histidines 1, 3, 11, and aspartic acid 76 coordinate the Zn2+ atom (1). An H1/3A double, an H11A, and a D76A single mutant of endostatin were not able to regress Lewis lung carcinoma. We conclude that the ability of endostatin to bind Zn2+ is essential for its antiangiogenic activity.
