[Obtaining of ScFv-CBD fusion protein and its application for affinity purification of recombinant human interferon alpha2b].
The gene of ScFv-CBD-fusion protein has been designed using the DNA sequences encoding of single-chain antibody (ScFv) against human interferon alpha2b (IFN-alpha2b) and cellulose-binding domain (CBD) from Clostridium thermocellum cellulosome. Biosynthesis of ScFv-CBD utilizing high-productive Escherichia coli system was carried out and the accumulation of target protein in bacterial inclusion bodies was shown. After the purification of the inclusion bodies and their subsequent in vitro refolding the soluble ScFv-CBD-fusion protein was directly immobilized on cellulose by bioaffinity coupling. The possibility to obtain the preparative quantities of ScFv-CBD in biologically-active form using different refolding schemes was accurately investigated in the paper. The general applicability of biologically immobilized ScFv-CBD-fusion proteins for affinity purification of recombinant IFN-alpha2b is shown.
Hil'chuk
P V
PV
Okuniev
O V
OV
Pavlova
M V
MV
Irodov
D M
DM
Horbatiuk
O B
OB
D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D002482:Cellulose / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D007128:Immunoglobulin Fragments / Q000235:genetics / Q000378:metabolism; D002479:Inclusion Bodies; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000276:immunology*; D010957:Plasmids; D016133:Polymerase Chain Reaction; D020673:Protein Renaturation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins
D000911:Antibodies, Monoclonal; D007128:Immunoglobulin Fragments; D000077190:Interferon alpha-2; D016898:Interferon-alpha; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C055966:immunoglobulin Fv; D002482:Cellulose