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· The
Abnormal Prion Protein is
 Rich in
β sheets
Protease
resistant
Detergent
insoluble
· Elastin has
VPGG sequence that imparts elasticity to the membrane
HYPOTHESIS
Is it possible to use a polymer
of both the elastin peptide and prion protein and generate an elastic
biopolymer?
Chemistry of the Solid Phase
Peptide Synthesis
Prusiner et al have been successful in using
a highly optimized Fmoc chemical technique
involving DCC/HOBt activation and an efficient
capping procedure with N-(2-Chlorobenzylooxycarbonyloxyl) succinimide. A single reverse phase purification step
gave a high yield of homogenous protein. This method helps to remove all
the side – products that accumulate during the various steps.
After synthesis, peptide sequence must be
confirmed by Edman’s degradation.
Controls – Use variants of the
VPGG sequence by introducing amino acids like Isoleucine.
Construct various mutant versions of prion protein with the inserted VPGG
repeats and compare them with the so called wild type polymer that has the
full length sequence of the prion protein.
Membrane Polymer obtained now
has to be subjected to various tests as follows
1. Secondary
Structure Determination by CD and FITR
2. Determine
Antigenic Nature of the Polymer –
In vivo tests in animals like mice will be the next step to
determine if these polymers are antigenic.
3. Determine
Physical Properties of the Membrane
Elasticity may be determined under physiological conditions.
Thermal stress will also be used to determine the protein’s
elasticity. Atomic force microscopy can also be used to determine the
elasticity.
4. Ability of
this Prion Polymer to induce conformational changes in the abnormal Prion
Protein
Incubation
of the normal cellular Isoform with the polymer and determine by Western
Blot and Proteinase K digestion assays.
APPLICATIONS
If
non-antigenic then this Biopolymer might find applications in medicine for
drug delivery, protect important tissues that are damaged, coat foreign
articles that might be needed to maintain the integrity of the body’s
organs.
References:
How Atomic Force Microscopy
works
Elastic Biomolecular
Machines; Scientific American January 1995; by Dan
Urry
Engineering the prion protein using
chemical synthesis; H.L.
Ball, D.S. King, F.E. Cohen, S.B.Prusiner, M.A.
Baldwin – J. Peptide Res, 2001, 58, 357 -
374
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