How do we determine the starting methionine in a polypeptide?

Question

From what I've learned from my first few college lectures, all proteins (or at least most of them, I think; correct me if I'm wrong) start with the amino acid methionine. My question is, when characterizing a certain polypeptide, how can we tell which methionine is the starting amino acid for that polypeptide chain, if the polypeptide contains more than one methionine?

Answer 1

First, if you are trying to figure this out from a DNA/mRNA sequence, you look for an open reading frame.  The best first guess is that the first methionine in that frame is the start codon.

Then, if you're looking in humans, you look for whether there is a favorable consensus sequence, i.e. a Kozak consensus sequence, named for it's discoverer, Marilyn Kozak.  Sometimes there are nonfunctional start codons upstream of that Kozak consenus sequence that are not actually used.  There are somewhat different consensus start sequences in more AT-rich organisms.

If you mean that you have the purified polypeptide on hand to characterize, then the starting amino acid will be at the amino terminus of the polypeptide.

Answer 2

If you're looking at the polypeptide, then the first one is the first one.

If you're looking at the mRNA, then the first AUG codes for the first methionine.

If you're looking at the DNA, then it's the first TAC that is coded for and not removed as an intron.

However, not all polypeptides end up beginning with methionine.  In most, the initial methionine is removed during processing in the ER.