ISSN:
1573-4943
Schlagwort(e):
Single-chain Fv
;
MOPC 315
;
anti-DNP
;
bifunctional fusion protein
;
recombinant antibody binding site
Quelle:
Springer Online Journal Archives 1860-2000
Thema:
Chemie und Pharmazie
Notizen:
Abstract The functional antigen binding region of antidinitrophenol mouse IgA myeloma MOPC 315 has been produced as a single-chain Fv (sFv) protein inE. coli. Recombinant 315 proteins included sFv alone, a bifunctional fusion protein with amino-terminal fragment B (FB) of staphylococcal protein A, and a two-chain 315 Fv fragment. Successful refolding of the 315 sFv required formation of disulfide bonds while the polypeptide was in a denatured state, as previously observed for the parent Fv fragment. Affinity-purified recombinant 315 proteins showed full recovery of specific activity, with values forK a,app of 1.5 to 2.2×106 M−1, equivalent to the parent 315 Fv fragment. As observed for natural 315 Fv, the sFv region of active FB-sFv315 fusion protein was resistant to pepsin treatment, whereas inactive protein was readily degraded. These experiments will allow the application of protein engineering to the 315 single-chain Fv; such studies can advance structure-function studies of antibody combining sites and lead to an improved understanding of single-chain Fv proteins.
Materialart:
Digitale Medien
URL:
http://dx.doi.org/10.1007/BF01025718
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