I am doing an assay for interaction with a substrate and I need a detergent that will interfere with the enzymatic activity as little as possible. What is the general recommendation in this case. E.g. I understand 2% SDS is bad, but what is better TRITON X100 or NP40 and at what concentration?
You are better off with non-ionic detergents. See https://www.gbiosciences.com/Non-Ionic-Detergents
from these I would go with octyl beta glucoside. Other sites will sell lauryl maltoside, which I have used in many enzyme refolding studies successfully. the detergent Z3-14 is excellent for membrane proteins but am not sure about enzymes. Finally, it is worth looking into amphipols, which are small organic molecules that do not produce suds and can replace most detergents for solubilization of highly insoluble proteins.
good luck.
If you are working with a soluble protein (i.e. not a membrane protein) in a plastic plate or cuvette, use a nonionic detergent at a concentration below its critical micellar concentration (CMC) to keep the protein from sticking to the plastic. I usually use Triton X-100, but you could also use Tween-20, NP40, Brij-35, or n-octylglucoside. If you are using a detergent that is subject to formation of peroxides, such as Triton X-100, then you should be careful about the age of the detergent and its storage conditions. I prefer to use 10% solutions provided in ampules under inert gas (sold by Thermo-Fisher as Surfact-Amps).
If you are working with a membrane protein, you will probably have to use a detergent at a concentration above its CMC to keep the protein in solution. You will have to experiment with detergents to find the best one. Some commonly used ones besides those already mentioned are CHAPS, n-octylglucoside, and dodecylmaltoside, but there are many others. Nonionic and zwitterionic detergents are preferable, but avoid anionic and cationic detergents because they tend to be denaturing. If the membrane protein binds the detergent tightly, you may be able to lower the detergent concentration below its CMC.
If the substrate is a lipid, there is a risk that the detergent will compete with the lipid for the substrate binding site, but you will have to use a detergent to keep the lipid in solution.
Could you elaborate which enzyme activity you are working with? Adam has given a detailed answer, and you have to decided which non-ionic detergent is available with your lab. Tween-20 is commonly used in ELISA for washing off loosely bound unwanted molecules that you may try.
I am using a small molecule to pull down proteins for proteomics (I am trying to identify the binding targets of that small molecule), so I do not know which enzymes will be bound, I do however know that some of them will probably be membrane proteins
There is, in my opinion, and to my knowledge - zero ways to predict this from theory alone. Sure avoid highly charged detergents that tenaciously interact with the peptide backbone, like SDS.. and probably sarcosine, but not necessarily, at rather low concentration for the latter. The defined, high CMC, 'monomeric' detergents may be useful - since you have a lot of control over them - a couple were mentioned. But I see no specific reason to favor a non-ionic over a zwitter ionic detergent - or one with a combination of features like Big-CHAP.
I've done enzymatic reactions successfully in the presence of Tween-20, Triton X-100, Nonidet P40 / IGEPAL CA630, and CHAPS. And most of the time is was happenstance which one was present.
The reason Tween-20 is used in ELISA and western blot protocols is because, again to my knowledge, although it is a "mild" detergent, it is thought to have a very protein-like surface binding profile on membranes and plastics, thus, it is itself considered a blocking agent.
Here's my favorite references on this subject:
Johnson, M. Detergents: Triton X-100, Tween-20, and More. labome.com 3, (2013).
Linke, D. Detergents: an overview. Methods in Enzymology 463, 603–617 (2009).
Seddon, A. M., Curnow, P. & Booth, P. J. Membrane proteins, lipids and detergents: Not just a soap opera. Biochimica et Biophysica Acta - Biomembranes 1666, 105–117 (2004).
And, of course - old-school versions of the CalBiochem catalog.
Good luck. Talk to you soon.
The use of detergent for membrane protein solubilisation, especially in a functional state, is not science but the blackest of arts. As a general rule anionic detergents (SDS, cholate...) denature, non-ionic detergents (C12E8, Triton...) may work and hardly anybody ever tries cationic detergents (CTAB, LDAO...). The latter is a mistake, I had very good results with CTAB. Also, look at the Anatrace catalog (www.anatrace.com), they have developed a lot of new detergents that look chemically more like (lyso-) phospholipid (Fos-choline or cymal series).
At any rate, you will have to do a lot of screening and optimisation to get this just right. I have covered membrane protein isolation in doi:10.1007/978-1-4419-7251-4_15.
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