doi:

DOI: 10.3724/SP.J.1206.2013.00091

Progress in Biochemistry and Biophysics (生物化学与生物物理进展) 2013/40:12 PP.1209-1219

Comparison of Autotransporter and Ice Nucleation Protein as Carrier Proteins for Antibody Display on The Cell Surface of Escherichia coli


Abstract:
Antibody surface display technology is critical for novel antibody screening and antibody affinity maturation. Currentlymost frequently used display methods are phage display and yeast display. Although Escherichia coli (E. coli) is easily cultured andgenetically manipulated, thus is potentially an excellent display host, the display technology based on E. colihas not been widely used.One of the problems is lack of efficient display of antibodies on the surface of E. coli. Many proteins have been tested as displaycarriers in outer membrane display in E. coli. Display systems based on autotransporter protein (AT) and ice nucleation protein (INP) isthe most extensively studied. Another problem is unstable survival rates of E. coli when antibody is displayed. In this study, wesystematically examined display level, antigen-binding affinity of displayed antibody and survival rate of E. coli using Ag43 β (β domainof Antigen43, an AT protein) and INPNC (fragment of N-terminal and C-terminal of INP) as carrier proteins and T7, lac, araBAD aspromoters for antibody expression. We found that the antigen-binding ability of the Ag43 β based display was superior to that of theINPNC based system. As expected, T7, lac and araBAD promoters drove high, medium and low expressions of antibody. The hostsurvival rate using T7 promoter was extremely low (INPNC: 0.0033%, Ag43 β: 0.02%, the host bearing araBAD promoter had thehighest survival rate (INPNC: 37.80%, Ag43 β: 90.23%), and the lac based system had a survival rate of 2.04% (INPNC) and 13.27%(Ag43 β). Balancing the antigen-binding abilities, antibody expression levels and survival rates, a system using Ag43 β as carrier proteinand lac as promoter is the best choice for antibody display on E. coli.

Key words:cell surface display,autotransporter,INP,scFv,survival rate

ReleaseDate:2015-04-18 09:14:50



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