It will depend on your protein(s). Some proteins (e.g. from thermophiles) are highly stable and resistant to denaturation and will not be denatured at room temperature and neither will some protein-protein interactions. In some cases, denaturation may even be incomplete upon heating, but that's rather rare. I would suggest that you try running a heated sample next to a non-heated one and look whether you see a difference in the band pattern

Hello,

I do it all the time, because my protein cannot be heated. You can add 10 mM DTT and 50 mM iodacetamide to get well defined protein bands on your gel without heating.

Of course you can! the unfold of the proteins will depend on much factors, in may experience is better to run the gel slowly rather than have a fast result. The resolution is better because you give time to the SDS present in the buffer mix well with your sample during the all migration. I hope it will be usefull for your. good luck!

If you feel that your proteins are sensitive to heat then you can heat between 40 - 60°C (for one hour) rather than heating at 95°C for 5 mins (the normal and usual heating procedure for denaturing proteins in SDS buffer).

In addition to answering your question, I would also like to discuss another issue. In fact, sometimes we don't know that what kind (heat stable or heat sensitive) of proteins we are having.

In my case, when I don't heat my samples then I have only one band (small molecular weight) in my proteins, however, when I heat my samples then I get several big bands. It looks to me that my proteins are heat sensitive.

I don't know if the proteins are not properly denatured (covered) by SDS then what will happen to them in the gel (other than their mobility). There are some researchers who say that if the proteins are not properly denatured (covered by SDS) then they can aggregate or cleave.

Well, the things are not clear, an expert advice is requested

It depends about the protein you use. Some proteins rapidly solubilize in presence of denature reagents such as SDS and/or beta mercapto ethanol, but others are resistent, for example, crystals of polyhedra derived from some insect virus. These crystals are resistent at least in presence of SDS, and therefore, you need to heat the sample to a correct unfolding. I hope it help you

Another purpose of the heating that you should take into account is that it denatures and deactivates proteases (relevant if your protein sample is derived from cell lysis). Proteases are active in SDS at room temperature. 

Hello Sanal!

I am currently performing WB without heating my samples. Instead I use an enzyme called Benzonase Nuclease. Here is the link to Sigma:

http://www.sigmaaldrich.com/catalog/product/sigma/e1014?lang=es®ion=AR

I am using it since my biggest issue is DNA present in my samples. I understand that you can also perform sonication but I've never used it. 

I have uploaded a picture of one WB where I used a heated sample vs a benzonase sample. Lane 1 : heated Lane 2: benzonase. Lane 3: heated (treated sample )Lane 4: benzonase (treated sample)

It works really great and saves a lot of time though it's quite expensive.