| Summary: | 碩士 === 大同大學 === 生物工程學系(所) === 97 === The inter-relationship of protein structure and function is of interest to
many researchers; however, formal academic study of denatured proteins,
arising e.g. from genetic mutations, has until now been limited. Of critical
significance are the degenerative processes that lead to the conversion of the
α-helix to the β-sheet, and the generation of amyloid plaques in for example:
Alzheimer’s disease, Parkinson’s disease and degenerative conditions
associated with prions.
Here a simple synthetic approach based on micro- contact imprinting
has been used to form imprints of the secondary structure of creatine kinase
(CK), denatured by treatment with SDS. The imprinted materials, formed with MMA and PEG400DMA, in a volume ratio of 5:95, have been
demonstrated to be able to separate denatured CK from its native from.
Using NaOH/trypsin, a template extraction-efficiency, i.e. denatured CK
removal from the film, of 90% was obtained. Subsequent re-binding of the
template, i.e. denatured CK, was typically 70%, (template solution 3.5 ×10 -7
mol, MIP surface area 1.69 cm2), with an imprinting efficiency of 9.7.
Evaluation of the imprinted polymers in non-competitive re-binding
experiments with native and denatured proteins, showed re-binding of 68.3%,
95.3%, 92.3% and 62.4% to the native forms of CK, IgG, HSA and myoglobin respectively; while the respective binding to denatured forms of
IgG, HSA and myoglobin was 94.1%, 92.7%, 71.7%. In a competitive binary
system, using the denatured forms of: IgG, HSA and myoglobin; the
selectivity was 98.7%, 96.8% and 63.8%, respectively.
The method used has been shown to be a successful approach for the
formation of high-affinity materials able to separate denatured CK from its
native form and from other denatured proteins. Such materials may find
future applications in practical sensing devices.
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