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Tagged recombinant protein has different mobility? - (Nov/28/2012 )

I did an expression of Adh gene in BL21 (DE3). I purified the protein using histidine tag column. Then, I run the total protein and also purified Adh protein in same SDS PAGE gel. BUT, the purified histidine tagged protein has a quite larger size than expected size. anyone can explain why this happen?Is it tagged recombinant protein has different mobility? anyone can explain this to me?

-An-nur Al-Irsyad-

It is relatively common for proteins not to migrate at their expected sizes. Adding a tag could easily alter the charge on the protein, and thereby alter how it runs. SDS PAGE gels are not totally based on protein size, with the SDS creating the all of the charge; PI and a few other factors also play a role.

-bob1-

SDS is detergent that give negative charge on protein-correct me if im wrong..
I though only native page rely on the protein charge, pI

-An-nur Al-Irsyad-

An-nur Al-Irsyad on Wed Jan 23 06:12:44 2013 said:


SDS is detergent that give negative charge on protein-correct me if im wrong..
I though only native page rely on the protein charge, pI
What you say is true. Native PAGE does only rely on charge and size, but SDS-PAGE also has some component of charge migration, despite the SDS.

-bob1-

this may give you some insight:
Attached File

-mdfenko-