Affinity and folding properties both influence the selection of antibodies with the selectively infective phage (SIP) methodology

FEBS Letters

Schwesinger, Falk¹  Pedrazzi, Graziella¹  Honegger, Annemarie¹  Krebber, Claus¹  Plückthun, Andreas¹ 

[1] Biochemisches Institut, Universität Zürich, Winterthurerstr. 190, CH-8057 Zürich, Switzerland

关键词: Selectively infective phage;    Single-chain Fv fragment;    Antibody engineering;    Antibody affinity;    Phage display;   

DOI  :  10.1016/S0014-5793(97)01143-5

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

We investigated which molecules are selected from a model library by the selectively infective phage (SIP) methodology. As a model system, we used the fluorescein binding single-chain Fv fragment FITC-E2, and from a 3D-model, we identified 11 residues potentially involved in hapten binding and mutated them individually to alanines. The binding constant of each mutant was determined by fluorescence titration, and each mutant was tested individually as well as in competitive SIP experiments for infectivity. After three rounds of SIP, only molecules with K_D values within a factor of 2 of the tightest binder remain, and among those, a mutant no longer carrying an unnecessary exposed tryptophan residue is preferentially selected. SIP is shown to select for the best overall properties of the displayed molecules, including folding behavior, stability and affinity.

【 授权许可】

Unknown