an in frame N-terminal deletion encompassing the start codon of a gene was identified. When I did western blot, I found 3 bands in the patient sample smaller than the size of the original protein and only one band of the actual size of the protein in the control. How can we explain and prove that the protein was translated using different starting codon representing the 3 different smaller bands in the patient sample?
Alternative start site, exon skipping (which may lead to frameshift), loss of protein modification site(s), and/or instability of the deleted protein are all possible explanations - and not mutually exclusive.
Dear Nicholas, thank you for your reply. I am asking if there is any known mechanism for prioritizing the alternate start codon. Why the translation didn't start with the first ATG codon after the deletion?! Instead it picked also two other ATG codons in the inside of the expected transcript
Is that fact, or interpretation? Do you have N-terminal protein sequence and/or mass spec, or are you extrapolating from three bands on a gel? Perhaps the later ATG have closer matches to Kozak consensus?
I have not done the MS yet. My deletion is in frame and the codon after the deletion is ATG so I expected only one band with smaller size than the full protein in the control. Interesting, I have found these 3 bands smaller than the control one. When I checked the transcript sequence after the deletion I found there are 3 start codons that can be used with the predicted MW coressponding the 3 band in western blot.
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