Overview of DNA Replication

Hi in this video we're gonna be talking about DNA replication. So DNA replication occurs differently um on the different strands of D. N. A. So remember we're dealing with the double helix. So there's two strands of D. N. A. It has to be unwound and replicated in order to have the two new strands of D. N. A. So we say that there are two template strands, there's the leading strand and the lagging strand and these are replicated differently. So the leading strand is replicated continuously adding nucleotides five prime 23 prime. This is the orientation on the new strand, the strand that's being synthesized. Okay. And it's always synthesized if you're talking about what's being synthesized, D. N. A. Is always synthesized five prime 23 prime. Um And on the lagging strand it um proceeds dis continuously adding nucleotides from five prime 23 prime as well. And I'll go back and correct this here but it's dis continuously here. And so um if the new synthesized strand is being made, five prime 23 prime, that means that the template strand. So what is being copied is read by the enzyme, three private five prime. So I feel like this is a very commonly a question on a test or a quiz. And people get confused because they're not sure which strand they're talking about. So synthesis happens five prime to three prime. So this is talking about the new strand that's being made. Whereas the sort of the reading happens three prime five prime. And that's on the template strand. So here's an image that I want to show and I really want you to look at this and make sure you understand how all these directions are happening. Okay, so D. N. A. Um is being unwound here, right? It started here and all of this is being unwound. And so the unwinding is occurring this way. Now when this portion say like let's just say that only this part right here was unwound. Well for the leading strand that works right, because it can just start replicating this way right on the three prime template. And it starts synthesizing from five prime on the new strand. And so if this part's unwound it's fine as the D. N. A. Is unwinding. It just keeps on replicating and it'll replicate all the way to the end. And that's the leading Australian. The lagging strand is different because it has to be read, it has to be synthesized in the same way. Five prime 23 prime. But in this case and the lagging strand, the D. N. A. Is anti parallel, meaning that the five prime is here on the template and the three prime is here. So it actually has to be replicated backwards right? Because this is the five prime and this is the three prime. But unfortunately we're still starting in the same place. D. N. A. Is still being unwound from here. But it can't start and go this way. It has to wait until the D. N. A. Is unwound to a certain point, say here where it starts replicating this way. So by the time it's replicated this some more D. N. A. Has been unwound and it can start here and it replicates until it gets to its other previous prag mint. Now as the D. N. A. Is unwinding, it can keep going, but it does this dis continuously as the D. N. A. Is unwinding right? Because not all the D. N. A. Is unwound this on the leading strand, it just sort of travels along with the unwinding enzyme, which we'll talk about until it gets to the end. But this one because it goes backwards essentially, it has to wait for this unwinding to occur. So it can start and actually replicate in this five or synthesizing this five prime 23 prime manner. And so it has to do this in the five prime 23 prime manner, Which is why the leading strand and the lagging strand are different because the orientation of each template is anti parallel. So one has to go one direction. The leading strand and the lagging strand has to replicate in the other direction. So if you ever get confused on you know what um direction everything is moving in, please feel free to come back to this image because there's always a question on, you know which direction things are occurring in and people get so confused on it and they usually miss the question not because they don't know, but because they're just unsure whether you know what's being synthesized in what direction. But I feel like this image is fairly clear and it's always a good resource to go back to. So with that, let's now move on.

Okay, so now let's talk about the steps of replication. The first step of replication involves unwinding the double helix. So remember the double helix is all double helix city and attached to each other. But in order for replication to occur, each one of those strands has to act as a template and it can't act as a template if it's bounced the other strand. So something has to come in and like break those hydrogen bonds in between the nucleotides. So that D. N. A. Double helix will be unwound into two single strands. So the enzyme or the protein that's responsible for this is called D. N. A. Hell a case and it attaches to D. N. A. And unwinds the double helix by breaking those hydrogen bonds. Now, once those hydrogen bonds are broken, you essentially have two single stranded pieces of D. N. A. Right? They're single stranded in some areas and there's still double helix where the hell a case hasn't reached yet. So it's not an entirely like single stranded piece of DNA. But it's most it's it's pretty much single stranded in that area. And so there are these special proteins called single strand binding proteins that come in and bind to those single strand regions. The regions that have been unwound already. Um And that prevents them from reforming right? Because these hydrogen bonds, they're gonna just spontaneously form, right, you don't need any capitalization of energy or anything like that. So if they're close to each other, they're just gonna bind back together unless something prevents them. So this is kind of just like a like a hat almost that gets put on that D. N. A. To protect it from binding to the other strand which is still close to it. And then as the DNA is unwound, what happens is a process called super coiling. Which you may have heard me talk about before. If not I'm going to talk about it later in this same topic. But super coiling is essentially where the D. N. A. Helix just get so wound up on each other. You can think of it as a rope if you had to rope wrapped around each other and you just keep spinning them right they're eventually going to super coil on themselves. They're going to roll up and get really tight. And that that winding is just going to keep those heli sees just being wound up really tight now as the D. N. A. Is being unwound in one area that actually results in that rope spinning that I'm talking about other sort of above. Um I don't want to use the word downstream but it's essentially what it is but it's at a different area ahead of that unwound ng so that portion of the D. N. A. Is gonna be unwound but it's not unwound yet. And so because it's not unwound yet and this portion being unwound, it's creating this tension in this D. N. A. Region. And that will result in this kind of rope coiling or what's called super coiling where it coils in on itself and there's special enzymes that actually get rid of that. Or else you know, the D. N. A. Would be able to be replicated because it would just be so tightly wound up. And those enzymes are called Topo Osama races. You may also see it as D. N. A. Dry race depending on the book you use, but it's the same thing. And so uh so here's an example of what happens. So here you have this double helix, right? And you have to unwind, you have to break these double hydrogen bonds in order to be able to unwind the double helix to leave them for um replication. So here you have your heel a case it's coming in and it's breaking these hydrogen bonds as it's breaking them. The single stranded binding proteins are coming in, binding to the single strands and preventing them from reforming these hydrogen bonds. That he'll a case just broke. And as it's doing this this region is becoming super coiled because you can't just unwind it here without it winding up here, if you don't believe me really, get some rope or some string or something and just try it out. Um And for yourself. And so these topo I saw Marie says come in and they prevent the D. N. A. From super coiling um on itself. So that's the first step is the unwinding. Then, once it's unwound replication can begin. And replication involves the use of a lot of important enzymes which we're gonna have to learn the names of. So the first thing is the paul three enzyme. Some of you may see it as the paul three hollow enzyme depending on your book. But essentially the paul three hollow enzyme consists of D. N. A, polymerase three and some other proteins. Accessory proteins that help it help it replicate. Now, D. N. A proliferates three is what replicates the D. N. A. Any of these accessory proteins that are found in this hollow enzyme are just accessory proteins right now. D. N. A polymerase three has to have these accessory enzymes because by itself it doesn't really like to attach to DNA for too long. It's like I got really other things to do. And so D. N. A polymerase three would add about 10, 12 nucleotides. And then it would fall off and be like, oh I'm ready to go do something else. But these accessory proteins help attach the plan raise to the D. N. A. For much longer so that it can replicate thousands upon thousands of nucleotides without falling off. So D. N. A polymerase does the replicating but without accessory proteins, it would be really inefficient at it now to start replication, you have to have D. N. A polymerase three. But D. N. A polymerase three actually cannot be the initiator, it can't be the first thing, it replicates it. But unless you tell it where to start replicating, it's just not gonna do its job. It has to be, you know, it's kind of like a four year old like giving a four year old a task, you have to make sure that it's doing it and starting in the right place, you have to make sure the four year old is going to stay at that topic. You know like you have to help them along on a plane race is very much like a four year old. So um it has to have these accessory proteins that are helping it stay on task. But it also has to have a special protein telling it where to start. And that protein is called a prime ace it's part of a prime a zone. Some of you may see it as prime a zone but essentially prima's ohm is just a big complex that contains the primates. Primates is the enzyme. And what primates does is it synthesizes RNA primers. Now the RNA primers bind to the region of the D. N. A. That's going to be replicated. So the primers bind there and it says hey D. N. A. Plymouth raise your four year old, you start here and so D. N. A polymerase is like there's where I start and it comes over and starts doing its thing and the accessory proteins make sure it stays on task. So D. N. A polymerase three recognizes the primers. It says this is where I'm supposed to start, it starts DNA replication now eventually um notice that these are RNA primers. These aren't DNA primers, right? But we're trying to replicate DNA not this mixture of D. N. A. R. N. A. So there's a different primaries, D. N. A polymerase one that comes in after, you know, summaries three is doing its thing sort of comes in behind it and says okay this is RNA. It shouldn't be RNA. So it takes out those RNA primers and makes them D. N. A. And that creates one long DNA strand, not some DNA with an RNA primer at the beginning. Now the primers are important for both the leading and lagging strand but you'll notice that the leading strand only needs one because it's continuous. And so it just continuously replicates. Where the lagging strand needs a lot more than 11 plus one plus plus plus plus because it needs one for every discontinuous fragment it makes. So let's look at this. So here we have D. N. A. It's unwinding right, here's the hell a case and it's breaking these D. N. A fragments as it goes. Now the leading strand. So it was started back here, right? The D. N. A. Was the double helix. And so it's some the hell a case. Probably started here, right? So it started unwinding d. n. a polymerase three came in and it started to replicate and it's replicating in this direction. And as long as the healer cases still unwinding things, it's going to go all the way to the end. The lagging strand, on the other hand, has to actually move in the opposite direction. So it's moving this way and replicating. So as um the hell a case is unwinding, it is starting to synthesize fragments. So here you have D. N. A polymerase three. So it's already copied these regions. So it probably um bound here and went this way and then it bound here and went this way and you can see that it will eventually bind somewhere here and go this way too. But um that is how the D. N. A polymerase three is replicating. But like I said, it's like a three year old, it needs accessory or four year old, it needs accessory proteins to keep it there. And it also needs an RNA primer. Now these RNA primers aren't very clear in this picture, but they're here essentially. So here's the primer. Here's this primer. So D. N. A polymerase probably started here and it's working on making this Okazaki fragment is what this is called. I'll introduce that word in a second and then once it's replicated this region, it's gonna start here where this primer is and replicate to here to hear where the other fragment meant. Then the primates will come in and make sure that this is replaced and made into a DNA or the DNA plum race, one will make sure that these RNA primers are turned into D. N. A. So uh this is an overview of how DNA replication works. So like I said before, I wanted to introduce some more terms for you. So the lagging strand is replicated dis continuously and this creates many D. N. A fragments and I use the word Okazaki fragment because that's what that fragments called, I want to produce it in a second. So the helix is unwound primates adds RNA primers. Um and then replication starts with those primers and it continues until it reached reaches the beginning of the strand or the previous fragment. Now this is occurring on the lagging strand, which is what we're talking about here. These fragments are called Okazaki fragments. And um once you have replication occurring on the lagging strand, you have a bunch of different Okazaki fragments after a while, right? Because you have just the discontinuous fragments that are being formed, but eventually you don't want a bunch of DNA fragments, you need an entire D. N. A. Strand being replicated. And so the enzyme responsible for connecting those fragments are called DNA is called DNA like and it joins the Okazaki fragments together which creates a single new replicated strand of D. N. A. So here we go. So this is the image that we use at the very very beginning and here we're too Okazaki fragments and you can see DNA legs has come in here and it's going to seal them together. So this I will add A D N A strands and will link them here. Or just like usually the DNA nucleotide that will link the two strands together and then it will jump here and do the same thing for this strand that will be made and it will jump here and do the same thing for this strand that will be made and so on and so forth until all of the Okazaki fragments have been connected together and become one DNA strand. So that is the overview of DNA replication or really just the detailed version of DNA replication. So hopefully that's clear with that. Let's not move on.

Okay, So now let's talk about DNA proof reading. So if you know anything about DNA replication know that it is replicated with extremely high fidelity. It very rarely makes an error. And the reason. So there's one error every 10 to the 10 nucleotides. And this is something that happens so quickly. Right? Nearly 1000 nucleotides are replicated per strand. So that means 2000 nucleotides per second. So not only is it very accurate, but it's occurring very fast. So it has this accuracy at very high speeds. And the reason it's so accurate is because DNA polymerase has proof reading abilities. So it's not always perfect, right? Sometimes it's going to make the wrong match. So it's going to pair an A. With A C. For instance or a G. With a T. Or so on and so forth. So that's called a DNA mismatch. And if the DNA mismatches made what the primaries does is it actually pause, pauses, goes back, cuts out that wrong base and replaces it. And so this type of activity is called X. A nucleus activity. And it's very important, you know that it occurs three prime to five prime. Know this because this is opposite, Right? So synthesis on the new strand occurs fried prime 23 prime. But proof reading occurs three prime, five prime. And it's important to understand these differences and the directional differences. And so the extra nucleus just means that it can cut out and mismatched nucleotide. So here we have an example of what this looks like. So we have D. N. A polymerase. It's going forth. It's just replicating itself and it's so happy and here has made an error, right? See doesn't go with T. It goes with T. And C. Goes with G. So this is a big error. So what it does is it actually pauses goes back cuts that nucleotide out that cuts that C. Out that's been paired wrong and using it 3 to 5 prime extra nucleus activity and then it replaces it and just keeps going. And so this proof reading ability is super important in giving D. N. A. Plymouth rates the high fidelity that we need to survive. If it didn't have this we would have a much higher error rate. We'd have a lot more mutations. Um A lot more health problems essentially. And we may not even be able to survive as humans without this proof reading ability. So with that let's not move on.

Which of the following proteins is responsible for unwinding the double stranded DNA?

• A. Topoissomerases
• B. Single-stranded binding proteins
• C. DNA helicase
• D. DNA polymerase III

DNA replication synthesizes DNA in which direction?

• A. 5' to 3'
• B. 3' to 5'
• C. Leading strand 5' to 3', lagging strand 3' to 5'
• D. Leading strand 3' to 5', lagging strand 5' to 3'

What would happen to DNA replication in DNA polymerase lost its 3' to 5' exonuclease activity?

• A. Replication would speed up
• B. Proofreading would stop and replication would stall
• C. Replication would only occur on the leading strand
• D. Replication would only occur on the lagging strand

Which of the following proteins is responsible for synthesizing RNA primers?

• A. Topoisomerases
• B. Single-stranded binding proteins
• C. Primase
• D. DNA polymerase III

The short DNA fragments created during lagging strand replication are called what?

• A. Primers
• B. Okazaki Fragments
• C. Replicates
• D. Exonucleases