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Current time:0:00Total duration:11:16

DNA repair 1

Video transcript

let's take a look at a segment of DNA that's in the process of being replicated and I want to focus in particular on the enzyme that replicates DNA and that enzyme is DNA polymerase actually there are a few different types of DNA polymerases on the one that we're looking at right now is DNA polymerase 3 so DNA polymerase 3 synthesizes new DNA and it also has the ability to proofread or kind of check the DNA that's putting together and make sure that there are no mistakes in it but before we get into that let's just orient ourselves and quickly summarize the diagram that we're looking at so this enzyme over here is DNA helicase that's the enzyme that unwinds the double-stranded DNA so that DNA polymerase can then come in and start replicating right over here you can see I drew the back one in a different color that's the RNA primer now let's just label the DNA strand that's being synthesized it's synthesized from five prime to three prime and actually the bottom strand of DNA is synthesized in the same time as the top strand but I just left that out of the drawing to keep things simple and let's say that the yellow bases represent the nitrogen base thymine let's say that the orange bases represent cytosine the green ones represent adenine and the blue ones represent guanine and so thymine and cytosine are the pyrimidines they are composed of a single ring structure so they're made up of one ring that has six sides to it and then adenine and guanine are the purines they are a double ring structure so they're composed of one ring with six sides to it and then that ring is attached to another ring that has five sides to it and actually the structures are a little bit more complex there are other atoms you know in it and there are some double bonds but we're just going to keep things simple for now and leave it at that and so let's get back to our D new this being replicated right over here I left a space I didn't put the nucleotide and I let's say that by accident instead of it being paired up with the proper base which is adenine it accidentally gets paired up with a guanine so that's a mistake and DNA polymerase three actually has the ability to sense if it made a mistake and if it does realize that it's going to go backwards and it's going to actually remove the incorrect base and replace it with a correct base so let's do that it's going to remove the incorrect base and replace it with the correct one of course you remember the nucleate the nitrogen base is attached to the sugar backbone and so this activity that I just described to you is called XO nuclease activity and so nuclease tells us that means the ability to remove a nucleotide and XO just going to underline that so XO tells us that it can remove a nucleotide but only from the end of a DNA strand so it was able to remove the nucleotide because it was at the end of a strand this is in contrast to endonuclease activity so an endonuclease can actually remove a nucleotide from the middle of a DNA strand so it would be able to remove I don't know a nucleotide from like right over here for example and just keep that in mind because we're going to come across some endonucleases as well anyway back to our exonuclease activity if we want to be more specific the exonuclease activity of DNA polymerase 3 is actually 3 prime to 5 prime exonuclease activity and the reason is called 3 part 3 prime to 5 prime exonuclease activity is because when DNA polymerase 3 makes that correction it has to move backwards in the 3 prime to 5 prime direction in order to do that there's another enzyme DNA polymerase one I'm just going to abbreviate polymerase with pol and DNA polymerase 1 also has excellent es activity and DNA polymerase wat is actually the enzyme that will remove the RNA primer at the end of replication and just as a side fact the exonuclease activity of DNA polymerase 1 is actually in the 5 prime to 3 prime direction so if you want you to just keep that in mind and so DNA polymerase 3 and DNA polymerase 1 are both able to repair or fix mistakes that happen during DNA replication and just to give you some perspective as to how often this occurs with and without repairs so normally we'll have a mistake happening in replication between 1 in 100,000 bases to 1 in 1 million bases that's normally the amount of mistakes that would occur but with their pure mechanisms of DNA polymerase 3 and DNA polymerase 1 this is reduced to a mistake that happens once in about 100 million bases and so they are very very effective at lowering the error rate in DNA replication so the next question I want to ask is what if this mistake over here was somehow not corrected during replication maybe there was something wrong with one of the enzymes something happened and that mistake was actually was actually sustained so let's take a look at that so here's a piece of DNA with our mistake incorporated into it and so before we discuss if this mistake can be corrected or not let's see what happens if this mistake is not corrected so right here we have our original DNA and we're replicating it and let's just say that this strand over here is the same as that strand let's say that the bottom strand in our original DNA is the same as this strand and so let's look first at the newly replicated DNA on the left so we have right over here finding bass and assuming DNA was replicated properly it's going to have an adenine complementary to it now let's take a look at the DNA on the right so on the bottom we had a guanine and it's going to be paired up hopefully with the correct base which is a cytosine now let's just quickly look back at our original DNA we were supposed to have a thymine with this complementary adenine and actually that's exactly what we got over here just going to circle it so this DNA is actually in the correct sequence but look at the DNA over here on the right this is not correct this is a mutation and this is an example of how mutations can occur if the DNA repair mechanisms are not working properly and so let's go back to our original question can we fix the original mistake so that this mutation does not occur and the answer that question is yes so fortunately our cells have what's called the mismatch repair mechanism and the mismatch repair mechanism is composed of number of proteins and the first thing these proteins are going to do is they're going to recognize if there's a problem and the reason that they're able to recognize the problem is that when you have a mismatch in DNA it tends to distort the sugar backbone a little bit and they're going to mark the area with a cup so they're going to cut the incorrect base or mark it with a cut the next thing is going to happen is in exonuclease is going to remove the incorrect nucleotide so we're going to remove the incorrect nucleotide the next step is one of the DNA polymerases is going to insert the correct nucleotide so we're going to pair our thymine up with adenine and the last step is a DNA ligase is going to connect the new nucleotide to the nucleotides on its sides and also to its complementary nucleotide on the other strand and I'm actually going to just correct that distorted sugar backbone and so here is our repaired DNA and just to clarify the mismatch repair mechanism that we're talking about here happens after application the repair is done by DNA polymerase 3 and DNA polymerase 1 that we discussed before that happens during replication or at the end of replication and so one thing you might be wondering is how does the mismatch repair mechanism know to distinguish between the original parental strand and the newly synthesized strand that has the mistake on it in other words how does it know which base over here is correct in our case that's the thymine and which one is incorrect in our case well it was a guanine and we know the answer to that question in bacteria so in bacteria the parental strand will have a deniz that are methylated so I'm just going to draw some methyl groups on all the adenine and that allows the mismatch for Pierce mechanism to kind of recognize and distinguish between the original strand that has the correct base on it and the new strand that has the incorrect base eyelet but we're not quite sure how the mismatch repair mechanism in eukaryotic cells and in other prokaryotic cells knows to distinguish between the Strand that has the correct nucleotide and the Strand that has the incorrect nucleotide