Hi all,
the Anti-Pyruvate Dehydrogenase E1-alpha subunit antibody from abcam normally running at 43kDa gives me an additional band at around 75kDa. Any ideas what this band represents? Could it be another subunit of the PDH complex?
Thanks!
My friend Safari
Let me give it a try
It is possible that the protein extraction process is faulty. Maybe the protein undergo some kind of modification process, thereby adding some additional weight to the protein, something we called translational modification of proteins which could be glycosylation or acetylation or any other form of modification. Assuming the molecular weight of the protein was lower, I would say the protein underwent some kind of proteolysis that cleaved off some amino acid residues leaving you with a lower kd protein. If you know exactly the type of posttranslational modification it underwent, you can treat the protein with hydrolytic enzyme that is tantamount to the modification. For example, you can use a glycosylase if you are sure it was glycosylation type of modification. The glycosylase will hydrolyze and cleave off the bond existing between the carbohydrate and your protein of interest
Another issue that might be the challenge is that the antibody may not be actually specific for that subunit of the enzyme. I could remember vividly that this complex enzyme in question also have E2 and E3 subunits (Pyruvate transacetylase and pyruvate decarboxylase if I am not wrong), maybe this antibody was not specific for that subunit. So, it is possible it is another subunit of the protein.
I hope this helps a little.
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