Ocr is unusual among proteins in its ability to form a transparent gel at high ammonium sulphate concentrations. This transition was investigated using a combination of spectroscopic, microscopic and rheological techniques. It occurs sharply at a concentration of 3.2M ammonium sulphate and is not observed with other types of salt. Rheological measurements showed that rather than precipitating under such conditions, ocr forms a weak viscoelastic gel. Far UV circular dichroism spectra reveal that ocr does not denature in the gel phase, while near UV CD spectra suggest the formation of long, helical structures. Well resolved fibrils were observed using atomic force microscopy. They were over 1µm in length and varied between 2.6nm to 10.4nm in height, corresponding to the thickness and length of the ocr dimer. Ocr is a highly charged protein (-56e at pH 8) and is shaped like a banana. We argue that it is stabilized in specifically aggregated structures at large salt concentrations by these physical properties. Electrostatic repulsions between proteins are screened by salts, allowing proteins to approach close enough to aggregate. The charge on ocr is high enough to resist such precipitation. However, at 3.2M ammonium sulphate we suggest that the salt molecules bridge neighbouring ocr dimers via hydrogen bonds, connecting amino acid carboxyl groups with the ammonium groups of the salt. The banana-shaped dimers stack on top of each other, forming long helical fibrils that intertwine into a semi flexible network.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:562085 |
| Date | January 2007 |
| Creators | Higham, Richard G. |
| Contributors | Dryden, David. : Poon, Wilson |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/1842/2569 |
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