Does the orientation of the VH and VL impact the binding properties of a ScFv when applied in a chimeric antigen receptor? Or are both configurations (VH-Linker-VL-CARbackbone vs. VL-Linker-VH-CARbackbone) equivalent?
Alejandro Martin
It depends. Most of the time it is best to leave the N-terminus of the VH free, as it usually makes the largest contribution to binding energetics (that's why most people use VH-linker-VL), but I once saw a case where a VL-linker-VH orientation worked better.
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Nagarjun Kasaraneni
I agree with alejandro martin, the design VH-linker-VL is mostly applied. The linker between VH and VL is also important for the design, mostly people use (G4S)3.
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