11_BL_20231030_WhisperAI_4.docx

So, now I would like to just give you a bit of an outlook on the eukaryotic systems that you will hear much more about in the classes that will follow. But how is DNA replication different when you look at it in prokaryotes versus eukaryotes? So first of all, the difference in the size of the genome...

So, now I would like to just give you a bit of an outlook on the eukaryotic systems that you will hear much more about in the classes that will follow. But how is DNA replication different when you look at it in prokaryotes versus eukaryotes? So first of all, the difference in the size of the genome is dramatic. Between E. coli, bacterium, and human, we are talking about a difference between millions and billions of base pairs. So we have three orders of magnitude increase in size. Our chromosomes are not a single chromosome but rather 23 linear chromosomes and so when you copy the DNA in the human DNA, you cannot start with a single origin of replication the way it happens in bacteria. So the bacterial chromosome, circular chromosome has one origin of replication where you have the replication bubble start and the replication forks will move with this DNA polymerase holoenzyme going in opposite directions until the two replication forks collide. Here you have to deal with multiple, 30,000 different origins at different places, so completely different beasts in eukaryotes. Eukaryotic DNA polymerases, they are now in the nucleus because eukaryotic organisms have a separate region where DNA is located, so-called nucleus. You have DNA polymerase alpha, that is called the initiator polymerase and this DNA polymerase actually will also synthesize the RNA primer. So instead of having a primer, a different enzyme, here you will have initiator DNA polymerase that synthesizes primer and starts DNA synthesis. And here we also can talk about this switching between the synthesis of primer and DNA and here you might remember that RNA duplexes or RNA DNA duplexes have different architectural features than duplexes of pure DNA. So RNA duplexes, for example, have this A form that is architecturally similar to the A form DNA. Then in eukaryotes you also have DNA polymerase beta, so alpha beta, and that one is responsible for DNA repair. And DNA polymerase delta is the one that is equivalent to DNA polymerase three in prokaryotes. That's the primary enzyme of DNA synthesis that is very, very fast. It can copy thousands and thousands of nucleotides at a very fast rate compared to these DNA repair polymerases. They don't have to be so efficient and so processive. So eukaryotic replication happens at 30,000 origins. Each origin contains a complex of six proteins and there is a specific place where the origin is recognized. Then helicase is recruited. That's the one that starts unwinding the DNA and then two different polymerases copy the genome. DNA polymerase alpha starts, synthesizes primer and extra 20 nucleotides. And then you have a very important concept, so-called polymerase switching. So you go from DNA polymerase alpha to DNA polymerase delta. This is referred to as very specific to the eukaryotic system. You have this polymerase switching. And then finally there is another unique aspect of eukaryotic genomes and that is the fact that they're frequently linear. So linear chromosomes means that they'll have ends. And you may remember that the initiation of DNA replication always happens with RNA molecule and that means that in this leading strand the beginning part of the DNA will be RNA. So it'll not be DNA. So this will be much more susceptible to damage and it'll be highly incorrect. So basically if you keep copying this DNA in every cycle of copying of linear DNA you will lose a little bit of this end because it will not be DNA but RNA. So how can that be fixed? Otherwise you have this erosion of DNA ends due to the basic aspect of DNA copying chemistry and mechanism. So these DNA ends are actually secured by repeat sequences that are not part of the fundamental genetic information. They're not responsible for synthesis of genes. These are so-called protective sequences called telomeres. They're repetitive sequences at the ends. They're called conservative repetitive sequences on chromosome ends. And the way these sequences protect the genome from being eroded upon successive cycles of copying is that for every cycle of replication there is a special enzyme that will simply extend those ends. So you copy the DNA that causes shrinkage because you start with RNA template but then the cell between divisions between copying of the DNA uses telomerase to extend these ends. Again it's copied and so forth. And this is very important process and if you don't have this enzyme you will cause damage to the DNA. This can cause early aging or it can cause cancer. There are many many bad effects if this system is not working properly. But what is especially interesting about this enzyme is that it has an RNA template, its own template for extension of DNA. So that means that the DNA can be extended even though there is no template for it and that's the key. That's why it can extend the ends without having to have complementary DNA because it has its own template. So this is called telomerase RNA piece. So RNA is used as a template for DNA synthesis. So basically you have this end of the DNA, you have a missing part and then this leading strand region in the 5' to 3' will be extended by the polymerase. So the extension again happens according to the usual chemistry of DNA synthesis and this is also something that I explained to you last time in 5' to 3' direction. You have extension then this polymerase will again bind. It's polymerase but it's called telomerase right and then it'll extend so then you have synthesis of this and so forth. So basically telomerase is an enzyme that uses its own RNA template to extend telomer ends. So with this I would like to conclude. This is the end of the discussion about DNA replication, recombination and repair and in the next hour we will start with discussion about transcription process. The second stage of this expression of genetic information from DNA ultimately to protein.