Lecture 3 - Translation & Gene Expression

Ajouté : 2026-05-14

[00:00:01]hi everyone welcome to our third lecture focusing on genetics specifically dna and how we're capable of being able to take transcripts of that dna to form mrna which is then translated to make proteins today we're going to be focusing on that process of translation and how we are capable of being able to regulate when we produce these proteins which is known as gene expression so two weeks ago we started looking at dna replication which is the process of how we duplicate our dna then last week we looked at specific segments of our dna which are known as genes and each gene has coding and non-coding regions and what we need to do is take a temporary working copy which is known as our transcript of that gene which we see in the form of messenger rna we went through the process of transcription where we formed this pre-mrna and then finally we took a look at how we go about processing this pre-mrna in order to produce the mrna that will be read and translated by the ribosomes through this process of splicing we looked at adding that five prime g cap and adding the three prime poly a tail in order to protect our coding sequence as well as how we can go about removing the non-coding regions which are known as introns and then splicing together the coding regions which you can see identified here in red remember we have this ability to be able to use alternative splicing methods where we can rearrange the order in which these exons are spliced together and that can give us a whole number of different possibilities for the sequence of mrna that we will produce so today we're picking up where that mrna has now left the nucleus through the nuclear pores it's now in the cytoplasm and it is going to be translated by a ribosome to produce our polypeptide chain so the mrna as we can see identified here has left the nucleus and is now in the cytoplasm and it has found a ribosome and it's the ribosome that's going to be responsible for producing the proteins each ribosome as you may recall is composed of both proteins as well as ribosomal rna and it's that ribosomal rna that's stored up in the nucleolus that dense region within the center of the nucleus when we need to make new ribosomes we utilize that ribosomal rna to form these subunits and then they leave the nucleus and in the cytoplasm they are assembled as our functioning ribosomes so you'll see here we have a large subunit and a small subunit and you may notice that there are these abbreviations here 60s and 40s those numbers identify the molecular size or molecular weight of those structures and the s or 60s and 40s stands for svedbergs which is a unit of measurement for measuring how fast a molecule moves when it is in a centrifuge so when they would take a sample of these ribosomes and centrifuge them which essentially means spinning them those that are larger would have a higher speed and those that are smaller would have a smaller speed and this is a very nice and simplistic model of the ribosomes but if we were to look at the molecular structure you can see that they are very complex remember they're made up of ribosomal rna as well as other ribosomal proteins now you'll notice here when assembled you can see that the large subunit has three specific binding sites and those are the a site the p site and the e site so remember mrna is going to be red five prime to three prime and so the five prime end of the mrna is going to be the first portion to enter the a site and so you can think of the a site as the arrival site and then the e site as the exit site the p site this is where the peptide bonds are going to be cleaved and that growing polypeptide will be transferred to the new amino acid that enters into the a site last week we spoke about trna and it's trna molecules that each contain a specific amino acid they are responsible for bringing in those brand new amino acids to the a sites to then continue building that complex polypeptide now last week i had spoken about the genetic code right we were looking at our mrna sequences and we were talking about how the ribosomes will read the mrna molecule in three base pair sequences and these three nucleotide sequences are known as codons and then when we looked at mutations last week we looked at what would the effect be if we say had a substitution to one of these three so you will have this chart accessible to you to reference it's not up to you to memorize the genetic code but you do need to understand how you can utilize it and so you'll see here you have on the left hand side here this would identify the first base of your codon up here at the top we would have the columns that would identify the second base of the codon so let's say your codon began with a the next c first codon would be a so we're in this row and then we find our second codon which would be c so you meet up at that square between the first and second codons okay so the row and the column where they meet up and then you will find that third base pair within that box so let's say it was aca well there you have it and aca codes 4 3 and e now remember if we had a substitution there that affected the third nucleotide in that sequence you can see that that would have zero impact on the overall amino acid that was coded for and that's an example of how we can have silent mutations of those third nucleotides and you can see here that that codon would end up coding for the exact same amino acid now each codon is made up of three different nucleotides remember our nucleotides are uracil cytosine adenine guanine because we are reading an mrna molecule so you look at your mrna molecule you read it in triplets and so say the first triplet which will usually be aug okay that means your first amino acid in sequence would be methionine and then so on now since each nucleotide within a codon could be one of four there's actually 64 possible codons that we can produce through rearranging those four different nucleotides in sequence now how many amino acids do we have in total we have 20. so you can see that if we have 64 possible codons and only 20 amino acids we're going to have multiple codons that will code for the same amino acid and that we identified by the fact that you could have multiple codons primarily if you change this third base that all code for the same amino acid and that is protective in nature because it means that our rna polymerase does not have to be perfect and so mistakes can still be made but they end up as silent mutations that don't affect the overall protein and function of that protein so last week i introduced you to the three types of rna we have messenger rna which we now know much about we have ribosomal rna which is responsible for producing the ribosomes and then we have trna and trnas are different molecules that each contain a specific anticodon in their genetic code and the amino acid that corresponds to the codon that they are going to be pairing up with so remember trna is ribonucleic acid so it is single stranded but you might be looking here and saying this looks a little bit double stranded this is something unique about the way in which the trna molecular code is set up last week we spoke about hairpin loops which are those regions at the termination points of transcription on our dna molecules well these are similar to those hairpin loops where you have a sequence here that then we'll want to complementary pair with another portion of that sequence if you look at the continuation here you'll see that there is all of these nucleotides in the middle of these two sequences but ultimately these are going to form complementary pairings but you can see here that from either side of that single stranded rna molecule there are going to be complementary pairings occurring and so the actual sequence of our trna molecules gives them this structure which kind of resembles a t now you'll see here they have what's known as an anticodon and the anticodon specifically pairs with the codon that we are trying to match up from the mrna molecule and you'll notice that the three prime end is responsible for binding the amino acid now in order for translation to take place the mrna molecule must bind to the smaller subunit when it does the other larger ribosomal subunit will then bind to it as well forming that complete ribosomal complex once the complex has been formed translation can then start the initiator trna is always going to be the trna that contains methionine can you remember what that codon was that codes for methionine it was aug aug is your start codon now there's also an enzyme known as peptidal transferase and peptidyl transferase is responsible for cleaving the peptide bonds that hold the amino acid to its trna here in the p site now our initiator trna is our trna that contains methionine so it will have the anticodon to match up with the codon aug now that mrna will be fed through to the p site until the next sequence is red and the new trna is paired up with the next codon in the a site we then require a new trna to move into that a site which will match up with the codon of the mrna in that a site peptidal transferase will then cleave the peptide bonds that hold methionine to the trna and the p site and will transfer it over to the trna in the a site you now have new peptide bonds that have been formed between the methionine and the new proline amino acid and you can see here that this is how our polypeptide chain is going to continue to grow now mrna is then going to shift further down so now this initiator trna that once had methionine bound to it it now moves into the e site so think of this as the exit site and the trna will dissociate from the mrna molecule and we now have the trna containing the proline and methionine existing in the p site and now this whole process continues a new trna will move into the a site and then peptidal transferase will cleave the bonds here from proline and we'll move the entire chain over to that new amino acid which is now in the a site and mrna will shift down okay and this is how translation occurs and so you'll see if you compare this to building say like a lego tower typically we would want to take new lego pieces and add them to the existing tower we wouldn't take the tower usually and go and stick it on top of the new piece this is how it works though with translation is that as this polypeptide continues to grow the peptidal transferase will always cleave the bonds within that p site and so the continually growing polypeptide is going to be then entirely transferred to the new amino acid that has just arrived at the a site now this process is going to continue until we finally reach a stop codon and remember we have a number of different stop codons we have one start codon aug but we have three different stop codons once the stop codon is reached the ribosomes will then dissociate from the mrna and the newly made polypeptide will be released into the cell now last week we talked about the idea of nonsense mutations and nonsense mutations were those that occurred where you would have a substitution that would code for a stop codon and so let's say we were at the fifth amino acid in sequence and it happened to be the codon and it happened to be coding for tyrosine so our codon was uac and let's say there was a substitution mutation where that c was substituted for a or g well in that case instead of coding for a new amino acid that would code for that protein to stop growing and ultimately would change the overall structure and probably make it non-functional so these stop codons exist normally in order to tell the ribosome to stop translation but we can also have nonsense mutations that can occur that can end up changing the codon to a stop codon which can end up yielding a non-functional protein now we've looked at this whole process of translation very simplistically right we looked at the one ribosome feeding the mrna and almost like paper into a shredder and the mrna is fed through and the polypeptide is grown but it's important to understand that multiple ribosomes might actually be responsible for transcribing a single mrna transcript remember in prokaryotes there's no nucleus and so we don't need to produce that primary transcript and they also don't contain introns and so they're able to transcribe very rapidly and so in prokaryotes transcription and translation almost occurs simultaneously where as the transcript is being formed the ribosome is also translating it and producing the protein and this is advantageous to them because they are quickly able to alter their regulation based on environmental changes and this is why they are capable of evolving so rapidly now in eukaryotes once translation is finished remember we have only created a simple polypeptide chain that's that primary protein structure well the protein is going to need to be folded into secondary but also tertiary structures and this will happen based on the arrangement of those amino acids in sequence now once that tertiary structure has been formed we might want to further modify that protein and specialize it by adding different fats or carbohydrates and so we can form lipoproteins and glycoproteins by either adding lipids or carbohydrates to our proteins within the golgi apparatus now glycoproteins we talked about antigens before right the markers on the surfaces of our cells well those antigens those are modified proteins they are glycoproteins so they're embedded proteins in our membranes that contain carbohydrate chains that act to identify specific cells such as our blood type for example now i want you to work through this example that i've provided you here during this week and then when we get into tutorial together that's where we're going to take this up together and work through this example so what you're going to be doing is taking a look here at your dna molecule we've identified here a template strand right which is the antisense and then you're also going to have the sense encoding strands you need to determine which one am i going to be using in order to produce my mrna transcript once you've done that you produce the mrna transcript so i want you to then take that mrna that you've transcribed and then break it into codons and once you've broken it into codons i then want you to use your genetic code and be able to determine the amino acid sequence so over the past two weeks we've taken a look at transcription and translation how the rna polymerase is capable of transcribing that pre-mrna molecule how we can then splice those exons together at the g5 prime cap and the three prime poly a tail in order to produce our mrna molecule and today we looked at when that mrna leaves the nucleus through the nuclear pores how we go about the process of translation and grow that polypeptide so now we're going to finish off the lecture this week taking a look at how we go about regulating when these genes are going to be translated now it's important that every cell can do this right we looked at recombination mutations that if they affect the regulatory regions that can impact how much of a protein maybe can be produced and this week we're going to start off looking at how prokaryotes specifically e coli bacteria are capable of regulating genes that are involved in the process of lactose metabolism now recall in eukaryotes we have transcription factors that must bind to the tata box which is a specific sequence of dna also known as the catbox as well located within the promoter region when the transcription factors bind our rna polymerase enzyme can then bind to that promoter region and form the transcription initiation complex in order for transcription to occur in prokaryotes specifically looking at the e coli bacteria today they have specific regions known as operators and the operators are capable of ultimately turning off transcription by having a repressor protein bind to that site and then also in prokaryotes we have the presence of an activator which can also make it easier for rna polymerase to bind to the promoter region and so today i'm going to use the analogy of a hot wheels race track and we're going to take a look at how the activator as well as the repressor are capable of regulating whether or not rna polymerase can bind and allow for transcription to take place now in order to illustrate gene regulation within the e coli bacteria we're looking at one specific series of genes that are located in sequence with one another that is known as an operon in this case here we're looking at the lac operon and the lac operon is made up of three separate genes each of which are responsible for producing functional proteins that assist the bacteria in metabolizing lactose okay so when rna polymerase is going to be transcribing this operon three functional proteins are going to be produced which are going to enable the bacteria to be able to take up and break down lactose now within the lac operon we have a promoter region so that should look fairly similar to what we've seen already in the eukaryotic cell in addition to the promoter before the transcription initiation site we have a region known as the operator and the operator is the site where the repressor protein will bind to and i want you to think of this repressor protein as the starting blocks all right this is like a gate and your rna polymerase this is your hot wheels car now if you've ever had one of those hot wheels race tracks what do we need to do in order to get power to the whole unit well we need to plug it in and so this cap binding protein here this is known as the activator and in order for anything to even remotely be able to occur we are going to have to have the power plugged into our hot wheels race track okay this cap binding protein is going to need to be bound to this activator site in order for there to be any possibility that we're going to transcribe these genes and we're going to make the proteins to break down lactose now you might be asking okay so okay i get the setup here we've got our dna as our race track we've got the cap binding protein that's acting as our plugging in power source the rna polymerase is our hot wheels race car and here we have some sort of starting block or starting gate okay so this is sitting in front of the car if we don't remove the gate it doesn't matter if there's power here the rna polymerase cannot transcribe the operon and it just so happens that the activator is regulated by whether or not glucose is available and the repressor is regulated by whether or not lactose is available so you'll see in the next two slides we have four potential scenarios that can exist and you're going to see in some of them the cat binding protein will be there or it won't and the repressor will be there or it won't so first off let's take a look at the cap binding protein the cap binding protein will bind to the dna when glucose is absent so let's think about this why is this e coli wanting to transcribe this operon well it's wanting to transcribe it to produce proteins that will help it take up and break down lactose and why would it want to break down lactose to break it into galactose and glucose so it can use the glucose to make atp right that's the whole point of all of this so think about this if glucose is present well there's no reason for this lac operon to be transcribed right if if glucose is present the bacteria can use that glucose in order to produce atp and so you'll see in these four scenarios whenever glucose is present the power won't be plugged in why we don't need this gene and as long as the power is not plugged in and the cat binding protein is not bound to the activator site the lac operon will not be transcribed all right so in any of these scenarios that's the first thing i want you to look for is the cat binding protein there or is it not if the cat binding protein is there it's because we don't have any glucose and so we're maybe going to need to use this lac operon to help break down lactose in order to get that glucose so in this first scenario here we can see glucose is absent and so the cap binding protein is bound to the activator site right we've plugged in our power because we don't have any glucose so we need somewhere to get it this might be a method that we can utilize in this case here glucose is available so we don't need the power plugged in because we don't need to break down lactose it doesn't matter if lactose is present or not if glucose is not present the cap binding protein will not bind to the activator site in this third scenario glucose is present again and so if glucose is present we don't need any lactose breakdown because we have a way in which we can access glucose and so again the cap binding protein is not bound to the activator site and this last scenario here glucose is absent so if it's absent we need to try to figure out where we're going to get some glucose so maybe we need to use this pathway and so here we go we're going to bind that cat binding protein to the activator site plugging in that power so glucose is dictating whether or not we're plugging in this machine what is then going to ultimately dictate whether or not the rna polymerase is going to end up transcribing is whether lactose is absent or present and in order for us to want to break down lactose we're going to have two criteria fulfilled glucose will need to be absent and lactose will need to be present right we need to have the lactose there if we're going to utilize this pathway we're going to produce these enzymes to help break it down okay so glucose needs to be absent and lactose must be present so in this first scenario here glucose is absent if it's not there we know we're plugging in the power but we have the starting gate here why do we have the starting gate here that's because lactose is absent the only way this repressor can come off of the operator's site is if lactose binds to specific binding sites on the repressor that makes sense right if we have lactose available we might want to break it down lactose is going to bind to the operator and the operator is going to come off but in this case here we don't have any lactose available so in this scenario lactose is absent and because lactose is not there it cannot bind to the repressor and so the starting blocks the repressor stays put so if the repressor's in the way rna polymerase cannot transcribe all right so the lac operon in this case is off let's move to the next scenario here the cap binding protein is not bound well if it's not bound that must mean that glucose is present glucose is available so we can utilize the freely available glucose and break that down to produce our atp we don't need the operon to be working but here look lactose was present how do we know that well because the operator's not there if lactose was absent well then we would see the repressor in position and now with glucose present we don't need the operon to be able to transcribe and so we don't even even initiate the process of getting an rna polymerase to come by the lac repressor is removed and the lac operon is completely non-functional now in our third scenario here let's take a look we do not have the power plugged in why don't we have the power plugged in because we have glucose available so we don't have that cap binding protein bound to the activator now in this case lactose is absent and because lactose is absent in this case the repressor is going to stay bound to the operator okay so our starting block is there but we don't have power anyway so it doesn't matter so the operon is off and you'll see between the first second and third all of those examples were situations in which the operon was off so what is the only condition under which the operon is going to be on and we're going to want transcription to occur it's when we don't have any glucose so the cap binding protein binds to the activation site and we have lots of lactose available and so lactose is going to bind to that operator the operator is going to come off and now that rna polymerase has the power it needs to go and your hot wheels car can then fly down the track right the rna polymerase now has nothing stopping it it's been activated and the repressor is gone and so now it can transcribe the lacz lacy and lac a genes which will ultimately lead to the production of the three functional proteins needed to break down the lactose so then the e coli bacteria is capable of utilizing the glucose broken down from the lactose to then produce its own atp now this was looking at how this one specific operon is regulated within the prokaryotes as we spoke about before eukaryotes have very complex ways of regulating their genes not only does transcription occur within the nucleus and then processing need to occur in order to remove the introns and splice together the axons we also have a number of different ways in which we can actually regulate the way in which specific genes will be expressed now because we have to fit all of our dna into that confined nucleus we need to condense the dna and we use those histone proteins okay it's almost like little little spindles that we wind the chromatin around to help condense it into those chromosomes and so there's going to be regions that are wound tightly and others that are wound more loosely and each cell is only capable of transcribing dna that is going to be accessible to the rna polymerase and so there's going to be different regions of chromosomes that are more condensed than others that are less condensed and so the highly condensed regions are not going to be transcribed as readily as those less condensed regions other ways in which we go about regulating genes we've already talked about alternate splicing we only have about 25 000 genes so 25 000 mrna transcripts that can be produced but remember we can splice together those exons in alternative arrangements and when we do that we produce well over a hundred thousand different mrna molecules and ultimately a hundred thousand different proteins now we can also add methyl groups to specific portions of our dna which actually act to prevent transcription and so that is known as a process known as gene silencing and this is a way in which environmental as well as dietary factors play a role in epigenetics that process by which we are capable of also up regulating or down regulating certain genes within our body something we haven't talked about yet but the fact that mrna transcripts all have different lifespans so those that are going to be more prone to degradation are going to have shorter life spans and those that will be less prone to degradation will be around longer so those with shorter lifespans won't be able to produce as many proteins as will those with say longer life spans and then lastly we looked at transcription factors this is one of the main ways in which our eukaryotic genes are regulated in order for the rna polymerase to bind to the promoter region transcription factors which are specific proteins they must bind to that tata box also known as a cat box which is that region within the promoter in order for the rna polymerase to bind and form the transcription initiation complex right without those transcription factors rna polymerase cannot simply transcribe that gene so those transcription factors are playing a huge role in how these genes are going to be regulated and we also have enhancer sequences within our eukaryotic cells that will also act similar to the activator sequence within the prokaryotes where we have specific regulatory proteins that will also need to bind ahead of the promoter in order to turn on transcription of that gene okay and that brings us to the end of our lecture for this week so we've gone through over three weeks looked at the process of dna replication and all the enzymes involved there where we're producing our replicated dna last week we started looking at transcription where we were looking at producing that mrna transcript and then finally how we can process that and this week we then finished off looking at how we take that temporary working copy that mrna and we translate it using the ribosomes to produce the proteins and then lastly looking at how we go about turning on and off various genes in our body so if you have any questions please don't hesitate to reach out otherwise i look forward to seeing you all in tutorial take care