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How to keep protein orientation during amide coupling?
- Bedo
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2 years 9 months ago - 2 years 9 months ago #1
by Bedo
How to keep protein orientation during amide coupling? was created by Bedo
Hello,
I want to immobilize the protein which has amine groups on the CM5 sensor chip which has dextran however I want to keep the orientation of a specific sequence intact so can I mix the protein with my analyte (which doesn't have amines) and can cover this specific sequence then introduce them after EDC / NHS. After that, I can wash with an acidic buffer to release the ligand/analyte interaction?
I want to immobilize the protein which has amine groups on the CM5 sensor chip which has dextran however I want to keep the orientation of a specific sequence intact so can I mix the protein with my analyte (which doesn't have amines) and can cover this specific sequence then introduce them after EDC / NHS. After that, I can wash with an acidic buffer to release the ligand/analyte interaction?
Last edit: 2 years 9 months ago by Bedo.
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- Arnoud
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2 years 9 months ago #2
by Arnoud
Replied by Arnoud on topic How to keep protein orientation during amide coupling?
In principle this should work. You must determine the amount of analyte needed to fully cover the ligand that you want to immobilize. In general, 10x KD will cover up to 90% ligand. Keep in mind that besides the amine on the Lysine the N-terminus of a protein is also a free amine. I tried to find an example but could not find it.
Arnoud
Arnoud
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