Previous studies have shown that it is possible to biotinylate proteins using the following method: Firstly a 15-amino acid acceptor peptide substrate (AP) is fused to the membrane protein. In presence of ATP and biotin, BirA catalyzes the site-specific biotinylation of AP, which can be read out by staining with streptavidin-fluorophore conjugates (Liu DS et al, PLoS One. 2013;8(2)).
Is there any similar method to conjugate a short ssDNA molecule to a membrane protein in living cells?
I would conjugate the oligo to streptavidin, and then have it bind to the biotinylated membrane protein. You can conjugate the oligo via its SH group ( which you can make or have made to have this), and add to maleimide activated streptavidin.
Or maybe during the biotinylation, in the PLos one procedure, you can feed the cells a biotin analog with the oligo attached. I haven't read the paper, so I don't know how the biotinylated protein is made.
Marcia: thanks for your answer. Do you know about any conjugation method where the biotinylation step (Liu DS et al, PLoS One. 2013;8(2)) can be substituted by a reaction with a modified oligonucleotide (purchased from IDT) ? I would like to have something like that: peptide+modified oligonucleotide+(enzyme or chemical)= peptide-oligonucleotide covalent bond.
Sorry, I don't. I don't know if this would exist. Of course, you can add things to a cysteine in a peptide.
You may be able to Maybe make a synthetic tRNA with it charged with a modified oligo? and then put a codon at the end unique to your synthetic tRNA? Maybe the oligo could be attached to a cysteine or lysine, and then use this to charge the new tRNA? The new tRNA can be made enzymatically. I think Tom Steitz Or Jim Wells made synthetic tRNAs.but I don't know if this can be done with intact cells. Ithink there would have to be a way for the cell to uptake the tRNA from the media.
Well, I hope this helps.
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