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19:10see the radioactivity, right? It's going to be 50/50. Okay, >> the the nitrogen that you put in that you tagged is either going to be there or it's not because the enzyme is either grabbing the left end or the right end. >> But if it is a concerted singlestep reaction then the nitrogen is always going to be there. The nitrogen that you put in on one end of this lysine is always going to be there. >> Okay. And that is what they found. They found that this is again this radioisotope measurement that they did. The other thing they did was nuclear magnetic resonance, the NMR, which is the stuff when we go to when we get, you know, um, our MRIs. Yes. It's the same technology. What they're using is that the nitrogen that they tagged has an odd
19:53number of nucleons in it, which means it's going to have a spin. And so you can measure that nuclear spin to figure out if >> your nitrogen is currently there. Okay? And they found that yep that nitrogen is definitely still there in our product which means that this is the asymmetric reaction because your nitrogen in the beginning was only on the left end. Yes. >> Right. And now the fact that it's 100% there all the time means this enzyme is always grabbing that left end. >> Right. >> Mhm. And so we've used the combination of observational tools and uh tracing tools to actually watch the evolution of the process >> to both show the stepwise and concerted
20:35versions and what happens. And because we're tagging the nitrogen, which is supposed to always be in the same spot, >> Yeah. >> each time, it's very obvious when you look at the tracing that in one of these pathways, it's always there. >> Yeah. Uh and this so it's a combination of these different levels of tooling that have allowed them to make this observation. >> Exactly. Yeah. And then they want to get into even deeper this enzyme. How does the enzyme actually work? >> Still not good enough. >> Still not good enough. Why is it different from the normal ones? >> Yes. >> Okay. So this class of enzymes they're calling it ornithine lysine arginine decaroxal oxidasis. The decaroxal oxidasis is key because it's both doing the decarboxilation and the oxidation in a single step.
21:16>> Um >> these are called Olados. >> Yeah, Olados. >> Olados is the uh the acronym. >> Yeah. And um they're from a class of proteins called PLPS, which use vitamin B6 as a co-actor. So the vitamin B6 is like helping along the enzyme even though it's not part of the active substrate. It's like in the back sort of like making sure that the enzyme shape is correct. Okay. Next, they used Alphafold. Okay. This is an AI tool that was developed by Google Google's deep mind to predict the 3D structure. >> Mhm. >> Before, let me just take a step back. Before we had AlphaFold, you had to like do all sorts of protein crystalallography, maybe like crym, all
21:59this other stuff. Now you can just plug it in to Alphafold and you'll get a really nice 3D structure. >> Plug it in. Plug it in. >> Right. And it's like it's it's super cheap to do. >> Mhm. Once you've got that, now we can figure out what is the difference between the ancestral olad and this particular oaddau. The difference is a tiny amino acid. Difference between tyrrosine and phenol alanine. Look at the structures of these two. They're