01929     2200205   4500001002100000005001500021035002000036245022100056700007600277700001900353250003800372260000900410300002500419084001500444082001500459008003900474650010300513520109200616990001501708INLIS00000000001769920210929081654  a0010-0721002231  aPurification Of Recombinant Peritrophic Membrane Proteins Of The Old World Screwworm Fly, Chrysomya Bezziana. /cPearson, Roger; Muharsini, Sri[Research Institute for Veterinary Science]Wijffels, Gene; Vuocolo, Tony.  aMuharsini, Sri[Research Institute for Veterinary Science]Wijffels, Gene  aVuocolo, Tony.  aJurnal Ilmu Ternak dan Veteriner.  c2000  aVol.5 (3): p.185-191  aARTVET2045  aARTVET2045210929                |          | |   4aScrewworm fly, Chrysomya bezziana, recombinant proteins, immobilized metal affinity chromatography  aTo evaluate the feasibility of vaccinating sheep against the Old World Screwworm fly Chrysomya bezziana several 
recombinant peritrophin proteins were expressed in either a denatured form in Escherichia coli or a native-like form in Pichia
pastoris cultures. Purification of the hexaHis tagged proteins was achieved by immobilized metal affinity chromatography.
Proteins purified under reducing conditions were refolded using a glutathione shuffle procedure. Purification of a glutathione-Stransferase
fusion
protein
was
attempted
using
glutathione
affinity
chromatography
in
conjunction
with
anion
exchange

chromatography.
The
authenticity
of
the
expressed
proteins
was
verified
by
amino
terminal
amino
acid
sequencing.

Carbohydrate
analysis
using
biotinylated
lectins
revealed
that
Cb-peritrophin-48
expressed
in
Pichia
pastoris
was
glycosylated

with
high
mannose-type
sugars.
Four
of
the
purified
recombinant
proteins
were
used
to
evaluate
their
protective
immunogenicity
in sheep against Chrysomya bezziana strike.  aARTVET2045