Hi Ali,

What makes this hard to answer, is that, yes, there are different tools out there that will segregate columns in an alignment between those that are better conserved, those that are less conserved. However, a major part of the reason why there are (many) different tools to do this kind of thing, is that they are designed with different purposes in mind, and thus "choose" their more/less conserved columns/regions in different ways.

Thus, without knowing why you want to put this selection of conserved regions - i.e. what are the statements you would like to make about (presumably the structure/function) this protein family (linking statements/inference of particular kinds of structure/function to different regions, perhaps) after identifying these regions, that you can't make without having done this analysis, I don't know which direction to point you in.

Working out/dissecting/identifying this is a crucial element of any bioinformatic analyses you do - and it's something that beginners to the field very often have trouble with. So please don't worry that you may be in this situation, it's normal!

My recommendation for you, is that you take some time to identify this i.e. identify the statements you're hoping to be able to make as a result of running your analysis e.g. "Due to their conservation, physicochemical properties, and the results of XXX prediction tool, these regions of the query protein are the ones most likely to be the transmembrane regions of the protein family" - once you've done that, let us know, and we may be better able to help you.

Just use ClustalW/T-Coffee/Muscle to align those sequences and then manually edit it if the conservered sequences in the literature are not in the same columns.