I would like to obtain a biotin-conjugated fusion protein. There are two methods reported in the literature. I wonder which one is more convenient and reliable?
The first method expressed purified AVI tag fusion protein from BL21 (DE3) host strain, and then biotinization kit was used to attach the biotinization protein.
The second method is to add biotin to the bacterial fluid when inducing the expression of target protein, so that the expression is the target protein conjugated with biotin, and then purify the biotinylated target protein through the purification kit.
Which method is more convenient and reliable?
Hello Jing Yang,
I think the first approach has much higher changes to work. Biotinylation will occur specifically on AviTag once you add biotin ligase, biotin and ATP.
On your second approach, I assume the biotinylation is performed chemically, which will decrease the specificity of your purification as many proteins will be biotinylated as well as your protein of interest. Also, it can inactivate your protein due to biotinylation of active sites and binding domains.
- Badges
- Science topic
- Similar topics
- Mathematical Sciences
- Statistics
- Probability
- Probability Theory
- Reliability