Biotinylation of a protein can cause a slight shift in its apparent molecular weight on SDS-PAGE, but the difference might not always be significant enough to result in two clearly separated bands. Whether they appear in the same band range depends on various factors:

In summary, biotinylated and non-biotinylated proteins may run in a similar range, but under certain conditions (e.g., highly biotinylated proteins, highly resolving gels), they could show up as distinct bands.@Elif Vera

Dear Elif Vera

MW of biotin is to small to detect the difference in MW

if you would like to detect it, you can try to add streptavidin and check the MW shift of biotinilated fraction. However it will allow you to disctiguish if a protein chain is biotinilated or not but not able to differentiate modo and multiple biotinilation.

good luck

Manuele

Hi Elif!

Yes, a biotinylated and non-biotinylated protein typically migrate in the same band range on SDS-PAGE, as the molecular weight difference due to biotinylation is usually minimal. Since biotin is a small molecule (~244 Da), the addition of a few biotin molecules results in only a slight weight increase, which is often undetectable, especially for larger proteins.