Bottom line: proteins like to move around – we call their different “poses” conformations and we call their shape-shifts conformational changes. Some parts of proteins like to move around more than others. And some proteins have lots of parts that really really like to move around. When regions move around so much that they have no real “set structure” we call them Intrinsically Disordered Regions (IDR’s). They’re “impossible” to see with structural biology techniques because they move around too much, canceling out their own signal, (and sometimes we actually have to chop them off in order to get proteins to crystallize) but that doesn’t mean there aren’t ways we can figure out stuff about them. And this stuff can be really cool! For example, proteins with lots of IDR’s can serve as scaffolds holding other proteins and/or nucleic acids together. A great example of this is GW182-family proteins (e.g. TNRC6A-C) helping connect the RISC complex (which is sequence-specifically bound to a target mRNA) to “generic” de-capping and de-tailing complexes that can repress whichever mRNA target RISC is bound to. Much more on this microRNA (miRNA)-mediated RNA interference (RNAi) here: http://bit.ly/microRNARNAi. IDR’s can also facilitate the formation of molecular condensates – kinda like gooey globs inside of cells that hold functionally-related things together to keep them from diffusing away from one another.

GW182 is basically like a big old spaghetti-like thing, but many proteins have regions within them that are disordered even though most of the protein has strong secondary structure (things like “set” α-helixes & β-strands – more here: https://bit.ly/proteinstructure ). Flexible linker regions often connect more “rock-hard” structural domains, and when proteins undergo shape-shifts (conformational changes), they often involve hinge-like motions taking place between the domains, with most of the “actual” changing happening in the linker regions. Even those “rock-hard” regions can change though, especially if they bind to something (such as Ago in a RISC changing shape when it binds to a target mRNA) or get post-translationally-modified (phosphorylated or something).

Sometimes you see the effects of something happening in one region of a protein somewhere else in a protein. We call these allosteric effects and they can involve shape-changes rippling through a protein from the “site of impact.” Such effects often come into play when talking about enzyme inhibitors that bind outside of the active site (so not competing with the thing (substrate) the enzyme can actually do things with) and when talking about membrane-spanning receptors that relay a signal from the outside to the inside.

Conformational changes can be subtle or dramatic and really really really subtle ones are taking place all the time as the protein “breathes” – local attractions between amino acids temporarily break and then reform. We know this sort of thing from techniques like HDX-MS (Hydrogen eXchange – Mass Spectrometry) which uses deuterium (heavy hydrogen) to label more flexible & dynamic regions of proteins. It’s able to sneak in during those breaths and swap out hydrogen for deuterium in the protein backbone.

Speaking of HDX-MS (much more on it here: http://bit.ly/hdxmassspec ), it’s one way that we can get info about regions that aren’t visible using structural biology techniques like x-ray crystallography (http://bit.ly/xraycrystallography2) & cryo-EM (https://bit.ly/cryoEMbumblyintro). Although the regions of the protein are physically there, you don’t see them in the signal so you don’t see them model. More on why here: https://bit.ly/crystalstructuremodels ). NMR can give you info about flexible things, but it only works for small proteins and you need a lot of it.

One technique that is sometimes used to study conformational changes is FRET labeling – for example, sticking a fluorophore on one part of a protein and a quencher on another. When those parts of the protein get close you’ll loose signal, but in a more open conformation you’ll see shining. More on fluorescence and FRET here: http://bit.ly/fretandfluorescence

X-ray crystallography & cryo-EM are super important, but they only show you a single “snapshot” of the protein (potentially more in cryo-EM if you have enough data to tease the different conformations apart). So it can be easy to think of proteins as “static things” but that’s far from true!

Sorry this post was weird. But I’m weird. And so are proteins. And that makes them awesome! (even if it makes them difficult to study!)

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