Suppose you had a piece of normal-sized paper, and you had some glue. Put glue all around the borders of the paper, and fold the piece of paper in half. You want the paper to stay glued together and perfectly folded in half – no bumps or creases.
The question: why wouldn’t you use the strongest glue possible?
The answer: because while you were folding it, every little mistake would be permanently fixed in place, so you wouldn’t actually get the right shape. Actually you don’t want the strongest glue, so that you can fold the paper in half and iron out the creases and bubbles once you have the basic shape down.
So now suppose you have a strand of RNA, and you get to choose its bases so that it folds exactly in half. You choose the bases, and you have the option of either choosing all G-U bonds (weak glue) or all G-C bonds (strong glue).
The question: why wouldn’t you use the strongest bonds possible?
The answer: because while it is folding in half with G-C bonds, every little mistake would be permanently fixed in place, so in the end you probably wouldn’t get the right shape. Actually you don’t want the strongest bonds, so that you can fold the RNA in half and it will be able to iron out the creases an bubbles once it gets the basic shape down.
How can I use this in the lab?
The basic thing you can take away from this idea is that the most-negative-free-energy design is not the best one. That design necessarily uses a lot of G’s and C’s (“very strong glue”), so while the RNA is folding and it inevitably folds a little bit wrong in some spot, it will have trouble unsticking at that spot in order to get to the shape that is actually the most stable.
So, very negative free energy is bad?
No. If your free energy isn’t very negative, then it is like not using glue at all – your RNA won’t stay in place even if it finds the right shape. The correct amount of free energy to use in the lab is likely a balance between making it sticky enough to stay together but not so sticky that it gets stuck the wrong way.
So, study up on those G-U competitions!
Chris - Fantastic Analogy! Brilliantly Articulated! Should be “Required Reading” for all new players before being allowed to enter the Lab!
The paper analogy is brilliant and instantly made sense!
This is great! I couldn’t have written it any better myself!
People have been wondering why the rules force you to use GU basepairs if they are so weak.
“An Analysis of G-U Base Pair Occurrence in Eukaryotic 5S rRNAs”
Here’s an article detailing the importance of G-U basepairs in RNA structures.
g-u base pairs are pretty important for RNA function, but they are very weak, so if you don’t make a rule forcing people to use them, the energy function by itself will not reward people for doing so. But it is an essential part of RNA design, knowing where one can and cannot put G-U base pairs.
Here’s a quote from the abstract
“In recent years, it has been shown that one of the most important structural elements in RNA is a wobble pair G-U. . . . The distribution of G-U pairs and the nature of adjacent bases suggests their possible role as a recognition site in interactions with other components of protein biosynthesis machinery.”
I feel that alan’s post here: https://docs.google.com/document/pub?..
should go here, just in case you already knew what potential energy was and wanted a more significantly-more-technical but significantly-less-handwavey version of the same reasoning.
Thank you. I’m new to this game but this explains a lot. I was only attempting to get the lowest energy and highest melting point.
I’ve also read that under certain conditions, Cytosine can mutate to Uracil spontaneously, turning what was once a very strong hold (perhaps the bond that is holding the structure together) into a very weak one. I don’t know how true it is, though.
@paramodic: yes, this is true. its called deamination–an amino group is lost from cytosine, turning it into uracil. this is a spontaneous reaction that occurs for sure in DNA, but im not sure if it also happens in RNA. in DNA its not a huge ordeal because there are specific ways of fixing it, but again–im not sure if RNA has a way of doing that.
Deamination occurs in both RNA and DNA. To my knowledge there is no way to detect, let alone repair these mutations.
I’m not sure that spontaneously deaminated cytidine residues are a big problem, though. Most RNAs turn over fairly quickly - even the genomic RNAs of viruses. Over the lifetime of the RNA, the mutation rate from spontatneous deamination is probably lower than the mutation rate caused by errors in transcription/replication. RNA polymerases are notoriously sloppy.
In RNA, cytidine deamination is usually deliberate. C -> U mutations are one of the more common changes made in RNA editing, and a very interesting protein called APOBEC3G is a cytidine deaminase that lethally mutates certain viruses.