AI Article Synopsis
- Aflatoxin B1 (AFB1) is a potent carcinogen found in food and feed, highlighting the need for effective detection methods through high-affinity antibodies.
- Previously developed scFv-M37, with 6 mutations, showed significantly improved binding to AFB1.
- The study found that reverting one of the mutations (VH-A110T) enhanced binding affinity further by improving the structure of the antibody, leading to more effective detection of AFB1.
Article Abstract
Aflatoxin B1 (AFB1) produced in Aspergillus flavus is a major hepatocarcinogen found in foods and feed. For effective immunological detection of AFB1 at low concentrations, the development of high affinity antibody for AFB1 is required. Previously, an affinity-maturated single-chain variable fragment containing 6 mutations (scFv-M37) was isolated from an artificial mutagenic library, which showed a 9-fold higher affinity than its wild type scFv. In this study, the effect of the 6 mutated residues on the affinity improvement was characterized using surface plasmon resonance analysis, which identified a deleterious mutation (VH-A110T) located on a framework region of the scFv-M37. The back mutation of VH-A110T resulted in a 3.2-fold affinity improvement, which was attributed to decrease of dissociation rate constant (kd) in interaction between AFB1 and the back mutant scFv. The biophysical analyses using circular dichroism and gel filtration revealed that the back mutation of VH-A110T caused a subtle conformational change of the scFv toward tighter binding to AFB1.
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| http://dx.doi.org/10.1016/j.foodchem.2016.04.085 | DOI Listing |
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