Mechanochemical Switches in Angiostatin.

Evidence is emerging that a mechanical tension can alter the functional states of proteins. It has been proved that a mechanical force can switch on the binding of PECAM-1 with a kinase (1), and the self-assembly of fibronectin into a fibrillar form by the exposure of cryptic binding sites (2). We previously reported that angiostatin modules mechanical unfolding was controlled by redox conditions (3). Here we report the first experimental evidence of a hierarchical coupling of redox and mechanical switches that controls the access to partially unfolded structures of biological significance in the multimodular protein angiostatin. We provide evidence that these structures can act as the antiangiogenic active forms of this protein, rather than the native fully oxidized and folded one as commonly considered so far. Among these structures we have identified a metastable intermediate that can account for unexplained data on the antiangiogenic activity of several angiostatin fragments (4) and for angiostatin binding to one of its receptors (5). The mechanochemical paradigm herein proposed could apply to other kringle fragments of multimodular proteins whose role in angiogenesis was proved, like prothrombin, apolipoprotein(a) and hepatocyte growth factor (6,7). 1. Osawa, M., Masuda, M., Kusano, K. & Fujiwara, K. Evidence for a role of platelet endothelial cell adhesion molecule-1 in endothelial cell mechanosignal transduction: is it a mechanoresponsive molecule? J. Cell. Biol. 158, 773-85 (2002). 2. Baneyx, G., Baugh, L. & Vogel, V. Fibronectin extension and unfolding within cell matrix fibrils controlled by cytoskeletal tension. Proc. Natl. Acad. Sci. USA 99, 5139-43 (2002). 3. Bustanji, Y. & Samorì, B. The mechanical properties of human angiostatin can be modulated by means of its disulfide bonds: A single-molecule force-spectroscopy study. Angew. Chem. Int. Ed. 41, 1546-1548 (2002). 4. Dettin, M. et al. Synthetic peptides derived from the angiostatin K4 domain inhibit endothelial cell migration. Chembiochem 4, 1238-42 (2003). 5. Veitonmaki, N. et al. Endothelial cell surface ATP synthase-triggered caspase-apoptotic pathway is essential for K1-5-induced antiangiogenesis. Cancer Res. 64, 3679-86 (2004). 6. Cao, Y., Cao, R. & Veitonmaki, N. Kringle structures and antiangiogenesis. Curr. Med. Chem. Anti-Canc. Agents 2, 667-81 (2002). 7. Sengupta, S. et al. Hepatocyte growth factor/scatter factor can induce angiogenesis independently of vascular endothelial growth factor. Arterioscler. Thromb. Vasc Biol 23, 69-75 (2003).