New neutron research at Institut Laue-Langevin (ILL) has revealed that the proteins making up silkworm silk have unexpected properties: effectively the proteins become more concentrated as they are diluted. The study published earlier this month in the RSC journal Soft Matter is a big step forward in understanding the amazing properties of silks and how to synthesise them.
There has long been interest in how a silk fibre is assembled from unspun protein precursors, but research has been hampered because it is difficult to obtain large enough quantities of 'native' samples (ie fresh from the worm) and few techniques are capable of investigating the structure of such large biological molecules.
Usually proteins are stable at a concentration of approximately 1mg/ml. As the concentration increases, say to 5-10mg/ml, proteins usually start to aggregate.
In this case, the scientists from Oxford and Lund found that the silk precursor proteins' behaviour is completely counterintuitive. Native concentration inside the worm can be up to 400mg/ml. "This is an extraordinarily high concentration for the proteins to remain stably dispersed throughout the solution," says Dr Cedric Dicko, a French biochemist working both in England and Sweden. "Even stranger, as the concentration drops the proteins begin to expand and flow, until they eventually clump together -- this is the reverse of what we'd expected."
At the native concentration the proteins form a compact helical structure with a radius of gyration of about 90nm; as they are diluted they unfold until they are 130nm in size. In the lab, the effect is a like a neat ball of string becoming unravelled into a big mess that ties itself in knots. However, the silkworms are able to control this process so that the proteins are spun into highly ordered silk filaments as they unfold and begin to flow. This surprising observation is a vital step towards understanding the liquid precursor, which is essential to synthesise silk and develop new materials with silk's desirable mechanical properties. The key lies in the detail, ie how stable is the precursor, and how is it is structurally and chemically modified during the spinning process?
The new insights also help to explain problems with previous studies based on 'regenerated' silk proteins. It is known that regenerated proteins (obtained by breaking down cocoon fibres with high salt concentrations) are of a smaller size and lower quality than native precursor proteins. Regenerated 'silks' are used because they may lead toward the commercialisation of spinning silk 'waste'. The new observations by the Oxford/Lund team allow the regenerated proteins to be compared with the native precursors. "It finally gives us a benchmark," says Dicko. "It shows that concentration affects the proteins' behaviour and therefore the type of material that can be produced, explaining the different results between research groups."
"Small angle neutron scattering was used to examine these large biological proteins in solution and was the only way to get this information," says Dr Phil Callow, one of ILL's instrument scientists. "There are also other advantages to using neutrons -- because they have no charge they do not cause any observable damage to biological samples. This means the same samples could subsequently be examined with the few other applicable methods."
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