Increased yield and activity of soluble single-chain antibody fragments by combining high-level expression and the Skp periplasmic chaperonin.
The success of recombinant antibody fragments as diagnostic reagents and therapeutic agents depends on the availability of sufficient functional material. We have produced a bacterial expression vector that combines high-level expression driven by a modified Shine-Dalgarno sequence with the periplasmic chaperonin Skp. Using this vector, we are able to obtain higher yields of soluble antibody fragments from cultures without the need for supplementation of the culture medium during expression. The fragments produced in the presence of the Skp show improved antigen binding activity compared to when the chaperonin is absent.
Mavrangelos
C
C
Child Health Research Institute, 72 King William Road, North Adelaide, South Australia, 5006, Australia.
Thiel
M
M
Adamson
P J
PJ
Millard
D J
DJ
Nobbs
S
S
Zola
H
H
Nicholson
I C
IC
D000818:Animals; D000937:Antigen-Antibody Reactions; D001426:Bacterial Proteins / Q000235:genetics; D018833:Chaperonins / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods*; D004268:DNA-Binding Proteins / Q000235:genetics*; D029968:Escherichia coli Proteins; D005434:Flow Cytometry; D005822:Genetic Vectors; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D016256:Lewis X Antigen / Q000276:immunology; D051379:Mice; D018832:Molecular Chaperones / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D007128:Immunoglobulin Fragments; D016256:Lewis X Antigen; D018832:Molecular Chaperones; D011993:Recombinant Fusion Proteins; C072721:Skp protein, E coli; D018833:Chaperonins