A structural glycoprotein has been extracted from bovine ligamentum nuchae, using 5M guanidine hydrochloride containing a disulphide bond reducing agent, and purified by preparative gel electrophoresis. The isolated material appeared to be monodisperse with a molecular weight of ~ 34 OOO as shown by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and by analytical ultracentrifugation. It contains 10% carbohydrate comprising mannose, N-acetyl-glucosamine, galactose and sialic acid in a 6:5:3:3 molar ratio. The glycoprotein has been assayed for peptidyl-lysine oxidase activity using 3H lysine-aortic elastin, prepared from 15- to 17-day-old chick embryos, as a substrate. In the absence of free lysine, the specific activity of the preparation over a 2h incubation was ~ 60 x 10<sup>4</sup> dpm/mg purified protein. Addition of 10mM lysine resulted in an ~ 50% decrease in the specific activity. Free lysine was shown to act as a substrate for the glycoprotein preparation as indicated by control experiments using 3H lysine in place of the aortic substrate. These results demonstrate that the glycoprotein exhibits a dual amine oxidase activity. In the presence of 0.27mM aminopropionitrile fumarate, a concentration which completely inhibits peptidyl-lysine oxidase activity in other lysyl oxidases, the glycoprotein preparation was inhibited by ~ 14%. In the absence of 5M guanidine hydrochloride and a reducing agent, the glycoprotein undergoes aggregation which in the presence of copper ions results in the formation of cylindrical tactoids, the diameter of which (11nm) corresponds closely to that of the fibrils which in the majority of connective tissue matrices constitute the microfibrillar component mainly associated with elastic fibres.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:751068 |
| Date | January 1981 |
| Creators | Ventrella, Giambattista |
| Contributors | Serafini-Fracassini, Augusto |
| Publisher | University of St Andrews |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/10023/14511 |
Page generated in 0.0361 seconds