Will endogenous and over-expressed protein level show different band size in western blot?

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Adam B Shapiro

It's possible that the endogenous protein has a post-translational modification, such as glycosylation, that the overexpressed protein lacks. Glycosylation in particular causes proteins to run at higher molecular weight on SDS-PAGE.

Pr Mei

Thank you for your reply. It is actually a glycoprotein but only two sites.

May I ask why over-expressed protein may not have glycosylation?

Didier Poncet

maybe your construct (a cDNA?) is missing one exon .... check with the genomic sequence...

Overexpressed protein may not be fully glycosylated if the cellular systems for glycosylation are overloaded, or if the overexpressed protein is not able to enter into the pathway due to misfolding or its site of expression.

wow many thanks!

I will definitely do some literature review haha.

Thank you for the answer.

The plasmids for over expression were confirmed by Sanger sequencing. should be fine.