Rotavirus is responsible for the death of many children annually, and current
vaccines have lower efficiency in developing countries. A reverse translated
consensus gene sequence of the rotavirus VP6 cloned into a pET-28a(+) plasmid
was used to transform BL21 and KRX Escherichia coli cells. Optimal expression of
soluble protein was induced in KRX cells by adding 0.05% L-rhamnose and 0.0001
M IPTG, with an incubation temperature of 25ºC for 6 h. VP6 was purified by
combining anion exchange chromatography followed by affinity chromatography.
Far-UV circular dichroism and intrinsic fluorescence were used as probes to assess
the native structure of VP6 and structural in the presence of a denaturant, high
sodium chloride concentrations and varying temperatures. The 0.2 M sodium
chloride had an impact on the VP6’s tertiary structure and also influenced the
proteins conformational changes as detected during thermal unfolding to 90ºC.
Although treatment with 3 M urea showed tertiary structural changes no secondary
structural loss occurred due to the presence of a denaturant. / Life Sciences / M. Sc. (Life Sciences)
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:unisa/oai:uir.unisa.ac.za:10500/23726 |
| Date | 12 1900 |
| Creators | Kgokolo, Samuel Maphalle |
| Contributors | Gildenhuys, Samantha, Parbhoo, Nishal |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Format | 1 online resource (xii, 99 leaves) ; illustrations (some color) |
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