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Cell Biology

Learn the toughest concepts covered in Cell Biology with step-by-step video tutorials and practice problems by world-class tutors

22. Techniques in Cell Biology

Isolation and Purification of Proteins



Protein Purification

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Hi in this video, I'm gonna be talking about the isolation and purification of proteins. So scientists need to study proteins for a variety of different reasons. But in order to do that, they have to obtain the protein for research. So they use a variety of techniques to do this, the first protein purification and this allows for the isolation of a single protein out of a big protein solution. So typically what you do is you can grow protein and bacterial cultures or you can grow protein in cells or you can just take cells that are growing and extract that protein. So you can extract it from bacteria cells, organisms, tissues, really anything. But then when you extract it you just have a mixture of this protein solution with all these proteins in it. So then you run it through a specialized machine and the specialized machine um fraction eight's the protein. So what that means is that you have this liquid and it runs it through a variety of different materials that separates it based on properties. So then if you run that protein liquid, that protein solution through this machine and it separates it by size for instance, then it alike. We caught these fractions or it sort of distributes these fractions, the small amount of liquid with only that size protein in it. So if you're looking for, you know, say you have this machine, this is the machine and it has the protein liquid going through and it's going to come out here, then you have all these tubes, these fractions and this one's gonna hold a five kg dalton, a 10 kg dalton 15 2025 30 35 40 kila dalton fraction or size of a protein. So when the five kila dalton proteins come out is gonna go in here. Then when the 10 killer Dalton proteins come out, they're gonna go in here and the same for the 15, the 20, the 25 and so on, so forth. So that would be an example by size fractionalization can also separate or fraction out the proteins in by charge or other properties whatever property the scientist wants. And in this process you can actually sort of when designing, you know, you need to purify this protein. You can actually tag your protein and to give them a certain property a certain size, a certain charge, a certain affinity for binding to something that makes them easier to purify. So let's say you have like you needed to separate out five proteins and they were all five kg daltons. It's gonna be hard to do this way because they would all go in this one too. But what we would do is you could tag those proteins with different things. Make one leave one the same. So then you'd have won five kill adult and 1 10 kill adult and 1 15 kill adult and so on and so forth. So that's protein purification. Getting out that one single protein from a solution. Then we have chromatography. And this allows for the separation of proteins again via certain properties. So there's a couple of different kinds you have column chromatography and this um runs that protein solution through this porous mixture has a bunch of different sized pores in it and that separates the proteins by size. You have affinity chromatography. This sorts them based on interactions with other proteins and then you have gel filtration chromatography. This also separates them based on size. So you can use different chromatography is for different reasons and they have different properties and all these different things I'm not talking about. But essentially the purpose of it is to separate out proteins. Then you have gel electrophoresis. This is a technique used to separate proteins based on a charge to mass ratio, so how big they are and how charged they are. But essentially once you get that purified protein, the whole purpose of this is to get a protein that's purified. So once you get it, the proteins can then be studied all by themselves. So in isolation and that's super important if you want to know the function of one protein and not the function of all of them. Super important. So this is an example of column chromatography. So here we have our column and you can see that there's some type of liquid here blue liquid. This is a protein solution has a ton of proteins in it. And what you see is that over time here's a clock. But over time there are proteins. Red one and a blue one will actually migrate. And so then you can collect the red protein and then you can replace this collection device and then collect the blue protein. So now you have one device with red protein in it and one device with, oh, that's black and blue protein in it. And that way you have those isolated proteins and these proteins can then be used to study something else. So with that let's now move on.

Which of the following methods allow for the isolation of a single protein?


Which of the following methods separates proteins by a charge to mass ratio?