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BIOL 2321_Microbiology for Science Majors - CH 07, 08 Video Lecture
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00:00 Um this is the start of the unit covering chapters 7789 and 10 and
00:08 six. So in that order, The uh chapter seven through uh 10
00:16 basically covering different aspects of uh bacterial procaryote, uh genetics. And we'll
00:25 with a bit of an overview really what Chapter 78 is about. And
00:28 very, very small portion of both these chapters. So make sure to
00:33 adhere to the, the pages you listed here and these are the pages
00:37 the 5th edition of the textbook. this will cover what uh reading that
00:44 , will just uh keep you on the material that we're, that's
00:49 gonna be covered in those two OK. So, uh topics to
00:55 there. First chapter seven, we're look at bacterial genomes, uh geno
01:00 , uh plasmids. Then chapter eight , chapter eight, a little bit
01:04 uh pro specific aspects of transcription, the uh R N A PLY and
01:10 function and then in translation um represent sequences which are specific to prokaryotes.
01:19 . And so kind of overall we're begin with like a little bit of
01:22 review of, of gene expression. I know you've likely had this
01:27 but that doesn't hurt just to go the basics of, of some of
01:31 terminology and kind of the overall uh of, of this, right?
01:35 very, very important um And especially , to know this, especially as
01:39 get into, you know, uh nine on horizontal gene transfer and on
01:45 chapter 10, especially chapter 10 with regulation, knowing and understanding what transcription
01:50 translation are because this manipulation of different in those processes is how we control
01:57 expression. And so um so very to have a good foundation of,
02:03 that. OK. Uh So now objectives uh for this section of Chapter
02:12 and eight. Uh So, as see listed here, you should be
02:16 to uh describe answer these uh different objectives. Uh Once you've completed this
02:25 module. OK. So uh let's with a little bit of a overview
02:32 starting with the, the terms genotype phenotype. OK. So here we
02:39 a picture of E coli bacterium And so, uh so you
02:45 of course, within the nucleoid remembers the uh the bacterial chromosome uh
02:51 And uh we can look at the genotype and phenotype in this context.
02:57 if we inoculate E coli into a uh a broth containing lactose sugar or
03:05 a plate, solid meat containing uh sugar we can see uh a reaction
03:12 taking place. OK. So if look at uh lactose broth, for
03:17 , uh e will give you a result. You see there, which
03:20 also a yellow, uh changing color , of the medium from yellow to
03:25 , a red to yellow rather indicating acidic drop in P H which is
03:30 of a bacter. That's fermenting a , you'll see a P H drop
03:34 then there's a P H indicator in medium that will turn turn to a
03:37 color indicating acidity. Uh That's an that this bacteria can ferment lactose.
03:44 . Uh A native uh result is on the right for comparison.
03:48 Similarly, on, on uh so , we can see eco has been
03:54 out and it's a lactose fermenter organism we saw from the Bronx results.
03:58 it will show as a, as reddish color on this particular type of
04:03 , which is uh called medium which for lactose fermentation, non lactose
04:09 pure colorless. OK. Again, due to P H change, changing
04:13 color of the P H indicator. . So what does all this
04:16 Well, it means this is one of how we can determine and these
04:21 are, are oftentimes used as a of the identification of E coli at
04:27 11 of one uh criteria for You, you would have several metabolic
04:34 . You would analyze and see what pattern is for all those. But
04:39 it's, it's reflective of the genetic of E coli, right? So
04:43 can kind of see what it is terms of in, in this
04:46 So, hence the term phenotype, we're what we're seeing here, we're
04:49 this change in this broth as cells from a red to yellow color indicating
04:56 fermenting the sugar. So it's so that's what we mean by
04:59 What is the, the actual characteristics appearance typically of an organism that's gonna
05:05 ? And that and that will be on what is the uh genetic capabilities
05:10 the organ? Because they both go in hand genotype phenotype. OK.
05:16 Now another example here is with blood . So with blood A uh there
05:21 bacteria that can when they're grown on medium or produce AAA clear zone around
05:29 colony where they're growing. Ok. that means, so blood auger contains
05:33 course blood and if there are bacteria are capable of lic red blood
05:38 Um and you'll see that as a zone where the enzyme that does that
05:43 hemolysin is diffused out of the cell is breaking up those her blood cells
05:48 it shows up as a clear zone the colonies. So again, just
05:51 another phenotype, right? But of , that what that action, that
05:56 we're seeing is due to a specific capability I E genes possessed by that
06:03 . OK. So a genotype of , translates into a phenotype.
06:08 just remember that it's not always about able to physically see things with your
06:13 like it's, it's of course, look at human and see that they
06:16 blue eyes and brown eyes or, uh you know, uh a,
06:20 different colored hair or what have you characteristic, physical characteristics we can see
06:26 their eyes, which are often due a genetic expression of genes that allows
06:32 that appearance. But remember that you thousands of chemical reactions going on inside
06:37 body that you don't see, but manifest themselves in different ways.
06:41 The point is the phenotype is a of the genotype, right? But
06:45 , you don't ever necessarily see all of the phenotype at, at
06:49 time. OK. And that relates gene expression as we'll learn.
06:55 um so another way to look at is here, here again is a
07:01 , this is using uh clubs and uh which is a good bacterium in
07:06 same group as E coli. And is a, a what's called a
07:10 ID test. And you see different , labeled uh glucose, lysine,
07:15 cetera. Uh We're focusing on the test and again, these are different
07:20 tests that depending on whether they're positive negative for these tests, you can
07:24 up with a profile and, and identification for the organism? Ok.
07:29 But anyway, so focusing on the uh test a positive result is a
07:34 fu color, purple, pink You see there is positive?
07:39 So there's a phenotype again, Appears pinkish. Ok. So what
07:43 that mean in terms of genotype? , how or how does that relate
07:47 genotype? And so of course, the appearance of the color, the
07:51 result is due to the enzyme. may, it, it, it
07:55 , it would possess to produce the result. So, what does that
07:59 do? Well, it now breaks urea which would be present in the
08:06 . Uh hydrolyzes it basically hydrolyzes it uh ammonium hydroxide which produces the P
08:15 change which gives the that purplish pink . So, um And he gives
08:24 co2 in the process. So, OK. So going further, let's
08:30 back now, this, of the reus enzyme is a protein.
08:34 . That can catalyze this reaction. how does that relate to genotype?
08:40 , the organism that would be positive the test would have the rease
08:44 right. So it would have the sequence that can has the information to
08:49 for the protein. Right? Here's gene expression, right? So,
08:53 a polymerase is part of this. you have the transcription translation process.
08:57 how we get from gene DNA eventually protein. In this example, the
09:03 ase enzyme. And so uh the A PLYM uh is all, is
09:08 a transcription making A R N A of the DNA. Then that M
09:15 N A resulting M R N A R N A or transcript is then
09:21 in the next phase using ribosomes. . And molecules called transfer R N
09:28 . So you have, so what obviously seeing here as you go on
09:31 road from DNA to protein in the is a lot of different R N
09:35 type molecules doing different functions carrying out translation. So uh the, the
09:42 of course come together to form that come together to form a large
09:48 and then collectively, they will translate form of the protein along with T
09:54 N A s of course. And that protein then will fall into a
09:58 that makes it active. And that's enzyme, it will carry out the
10:04 of break of hydrolyzing dia to ammonium . So that's how this all comes
10:10 . So you're seeing initially, we a positive result, the pink
10:13 What does that mean that result is to a specific enzyme? The bacterium
10:20 it has the enzyme because it has gene for the enzyme that codes for
10:24 . OK. And um genes that expressed can, can, will,
10:29 produce a particular phenotype. OK. uh so lastly is going through kind
10:37 how this all ties together. And is uh the, the process,
10:42 ? The central dogma. That's one those unifying concepts in biology.
10:49 There's several of those as you've, gone through intro bio uh courses.
10:55 you've gone through a number of different concepts. Uh Probably one of the
11:01 ones, if not the most important is evolution is a unifying concept.
11:05 But so too is a central dogma it's universal uh all living uh beings
11:12 this uh DNA to R N A , how genes are expressed,
11:18 And how they all are also OK. Um So kind of putting
11:25 all together then or sum and summarizing . So we have the transcription
11:29 So uh a chromosome will contain, course, thousands of, of
11:36 the oxy the A G C T a particular sequence. Um Those uh
11:44 we call genes can be both protein genes. Um There may be nonprotein
11:51 genes that set this, that express at the R N A. The
11:54 N A is a product molecule. But there is DNA that's doesn't code
11:58 anything and is typically usually involved in type of regulatory control. OK.
12:05 for those coding genes, then um the, the DNA is the think
12:13 it as the um there's a it's a storehouse of information,
12:19 And um you retrieve information from DNA the form of R N A
12:27 messenger R N A S that are of specific genes in the chromosome.
12:35 . And uh those M R M N A s are produced Under conditions
12:41 they are needed. OK. This all driven by which we'll learn in
12:45 10, you know, which genes being expressed are all driven by.
12:49 are the needs of the organism, . For bacterium in the environment or
12:55 your gut, you know, there's sorts of signals in uh external,
13:00 um that are um that are um if you will by the cell to
13:10 which genes are expressed or not right? Presumably those being expressed or
13:15 that are needed at that particular time do whatever functions are necessary.
13:19 Which could be a, a lots different things, everything from uh from
13:26 taking in a nutrient and, and able to utilize it to metabolize it
13:31 maybe dealing with some sort of a , maybe some kind of osmotic stress
13:35 other that it must deal with. there's a, a physical temperature change
13:38 something. So these are all environmental that cells deal with and they'll deal
13:43 typically through the functioning of different Uh But of course, those proteins
13:50 made until the genes for them are . OK. So uh no,
13:56 one organism on the planet Earth ever expressing all of its genes at one
14:00 . OK. It's what is needed that there are some genes that are
14:06 , need to be expressed. All time because you have functions that basically
14:08 always going on and there's others that , are expressed only at certain times
14:13 they are needed, right? So , it fits the spans the whole
14:17 . OK? You have genes you expressed since you were a zygote and
14:23 plus 10 days. You don't need genes anymore because you've already, you're
14:28 fully developed human, but you needed back then to carry out the proper
14:33 of development. So, you every organism typically will have genes that
14:38 that category. But you know, course, can be more complicated depending
14:42 the type of organism you are, human versus a bacterium. So,
14:46 even between that comparison, there are in common. OK. So
14:50 back to transcription, translation. So transcription provides the, the, the
14:55 N A copies with which the cell translate into proteins. OK. Um
15:00 translation process involves um the genetic the transfer R N A molecules for
15:07 R N A molecules. Um And you see the DNA template here of
15:13 portion of a DNA template of uh this chromosome. And so we also
15:18 the term sense and anti cents coding . I'll mention that briefly here in
15:22 second. So um so the antisense is uh of DNA is what is
15:31 by R N A plea to produce sense R N A strand, a
15:36 M R N A and that sense can be translated into protein.
15:41 And so the elements of a transcript have a ribosome binding site uh which
15:46 allow ribosomes to bind and then begin translate. And this you get a
15:51 on. So there's punctuation marks in transcript much like a sentence. You
15:56 , when you're reading a book, know, there's obviously hundreds and thousands
16:00 sentences in the book. But you the structure of how it works.
16:04 first letter of the sentence is capitalized there's a period at the end.
16:07 , you know, there's certain grammatical type elements that define a sentence the
16:13 in transcription. So you can define transcript and be able to translate it
16:17 looking for certain things. And one those, of course is what we
16:20 a star codon that begins the the the message uh and a stop codon
16:26 ends it, right? And coons between, right, that, that
16:30 what the protein will be. So those are the elements that,
16:34 are, that are utilized to produce a functional polypeptide. OK. And
16:41 is where the course a genetic code comes in, right. And um
16:46 and so again, back to these , sense uh antisense coding, non
16:50 , just briefly mentioned that that's important , to know that be familiar with
16:55 comfortable with those terms sense, antisense encoding and what they mean, especially
17:00 we get into viruses because of different of viruses. Um depending on the
17:07 , the genome can be a sensor strand and that can have different consequences
17:12 terms of their reproductive cycle. Nevertheless, so taking the previous sequence
17:18 have here and just putting it here's the scent strand and anti cent
17:25 and it's labeled. Ok. always remember that the scent strand is
17:30 coating strand, right? It contains information that will produce the protein.
17:35 . The antisense strand is a copy that and it's, and it's the
17:39 , the antisense strand is the template will be copied. OK. You're
17:45 , well, if the sense car the essential information, then why are
17:48 copying a template strand? Simple explanation that, right? And it has
17:53 do with the base parent rules of acids. OK. So here would
17:59 the M R N A that would copied from the antisense strand,
18:03 So it's complementary base brain, C O G. Uh remember R
18:08 A s don't have thymine, they cells, right? So uh any
18:13 you see an ad in DNA, pairing will be with, as you
18:21 right here, uh not with the . So M R A s are
18:26 have T S in them. No means they're gonna have A S US
18:29 S and CS. OK. So , so the M R N A
18:34 is um represents a copy of that or template strand, OK. And
18:43 see the complementary base from G C T A, et cetera.
18:48 So then that M R N A represents a sense of strength,
18:52 So whether it's you can have a , DNA, which you see
18:57 right? You can have DNA R A pair, you can have R
19:01 A R N A, OK? so it's always gonna be uh complimentary
19:08 each other, right? So you the five prime three prime cent
19:15 the complementary strand is three prime, prime, right? So those rules
19:19 as well. So this here you five prime, three prime,
19:24 So they're always complimentary to each So the M R N A,
19:27 you look at closer, look at , it's a sense strand and you
19:33 it to the DNA sens strand, see that they're identical except for teas
19:40 substituted with OK. So what this the point here is that the M
19:46 N A is a copy identical copy the sensor strand, right? The
19:49 strand contains the coding information. So M R N A is now representative
19:55 that coding information right in an R A form. And that can be
19:59 . OK. And that's what happens you copy the antisense strand because it's
20:05 we'll produce, then by copying the sensor template strand, we essentially produce
20:10 copy of DNA sense strand which contains essential coding information, right? And
20:15 we look at the punctuation, There's your a your um excuse
20:22 there is your A U G. right. That's your start coat
20:26 That's actually a meine met for Uh This would be a stop coat
20:32 that actually that yeah, that would a stop coat on. And then
20:38 have code uh amino acids in right? So remember that the
20:42 the coons are the three base ReNu sequences that code for an amino
20:48 And that's, and that, and is deduced from the um genetic code
20:53 . So you look up G C in a genetic code table and we
20:57 tell you a specific amino acid, where the T R N A s
21:00 in. That's their role is is to match up with the
21:05 The transfer R A has a anti that matches up with the proper codon
21:10 brings the amino acid with it. that's how you build a polypeptide chain
21:15 Rizo moves along the M R N and transfer R N A is coming
21:20 at a time that match up with proper codons, bringing the proper amino
21:24 . OK. So these are things should be familiar with. OK.
21:28 , and have learned in bio. right. So uh so that's how
21:34 , so it works out. um so uh and of course changes
21:40 be made in the sequence. You have mutations that can occur in DNA
21:46 that can alter the polypeptide sequence. . Most of the time those changes
21:52 usually either uh silent. In other , there's no change at all or
21:58 lethal. Right? Because the changes detrimental, but there are, of
22:02 , times when they can be Right? And that's when mutational changes
22:06 occur. Ok. So, let's talk a little about proo
22:14 right? So there's different hierarchies if will, we can look at it
22:17 terms of the genome, the transcriptome the proteome. OK. And it
22:22 translates to uh DNA R N A , right? The central dogma,
22:26 genome of course is the is the of DNA uh A cell has and
22:31 can include both the chromosome and for , it may include smaller circular pieces
22:37 DNA called plasmas. OK. So genome is all the DNA with a
22:43 . And for bacteria that will of course, the chromosome can also
22:46 the plasmids. OK. So remember pros or haploid and OK, they
23:07 one chromosome, they are diploid like that have two copies of each
23:11 So they have 11 chromosome, they're . Uh And again, it may
23:16 include additional plasmids in addition to the , the transcript to of course,
23:22 the R N A is, it's transcripts that are produced at any given
23:25 . OK. And uh transcripts was , do not hang around indefinitely.
23:35 actually are go away after, in , they go away a actually rather
23:41 uh in, in minutes, they're around and they go away.
23:45 the cell needs to have more, can always do more transcription. Uh
23:48 they, but, but they don't around all uh indefinitely because as long
23:52 they are, are hanging around, will be translated and remember, bacteria
23:56 be very efficient and only express genes need to and for as long as
24:00 need to. And so uh transcripts dimensionally just disappear on their own after
24:05 few minutes. Uh But if they can produce more and, and
24:09 more transcripts means it can produce more . And that's what the protein
24:13 OK? With all the different proteins expressed at a given time.
24:18 So remember, it's only gonna be here, right? Portions of the
24:22 are being transcribed and then those are translated into protein, right? So
24:26 is no cell they will ever ever be expressing every single gene,
24:32 ? It's all about what's what's what's needed at any given time.
24:36 , genome memorization. So as I , uh most are, are
24:41 You can have a phenomenon called partial . We'll talk about that in Chapter
24:47 . Uh But, but, but a rule of bacteria, archaea do
24:51 possess two complete chromosomes. It's one uh size order of 500,000 to uh
25:01 . The average size I'd say is a million base pairs. E coli
25:04 like, I think 4.5 million base . Um Most, most bacteria archaea
25:12 the mold of uh you know, circular chromosome. Uh But there may
25:18 some that have plasmas as well. . All that's gonna comprise the
25:22 right? And we'll talk more about here shortly. Um OK. The
25:30 . So for comparative purposes only, going to show you the eukaryote um
25:36 and how its genes are organized. you will not be tested on
25:41 Uh on the exam, you will tested on eukaryote gene structure, but
25:45 will be of course for pro Caro structure. So the procaryotes, they
25:49 um uh very efficient um compact Uh They have uh On average,
25:59 translates to about 3000 genes I would is probably the average for most
26:04 3-4,000 genes uh that they possess um the single gene versus operant. So
26:11 structure is particular to pro caros. . Um They do have some genes
26:17 are just single genes. OK? a large number of them are organized
26:22 an operon, right? And OPERON basically, you know, one of
26:25 key key elements of any gene structure the term you see they called
26:31 OK. This is true for any you Caro the promoter is key because
26:38 what orients, it's what defines the of the gene. OK. And
26:45 so the promoter look uh can also a control element. And um the
26:54 uh the opera structure is utilizes one in multiple genes. OK. And
27:02 we'll look at that structure here in minute uh control sequences. So as
27:05 mentioned, a promoter, there can uh uh other sequences that bind regul
27:11 proteins that are involved in control. . Terminology, the cyst cyst is
27:16 a gene. OK. So mono structure is one promoter, one
27:22 a polycystic is one promoter, multiple . That's what a operon structure is
27:28 is is polycystic. OK. Um , Operon and Regulon, we'll talk
27:35 talk about Regulon as well. Regulon basically uh collections of operon that pertain
27:42 a similar metabolism. Right? I'll elaborate on that shortly. So
27:48 comparison, then the EU structure right , can be somewhat complex. And
27:53 there are control elements, uh those are close by the gene which are
28:00 proximal and those far away. Typically are things like enhancer sequences that serve
28:06 increase the levels of gene expression. . So um we'll talk about In
28:13 10, about what we refer to basal gene expression, which is basically
28:16 very low level expression. And so typically need that amped up to produce
28:24 levels of protein to do something. that's where different control elements come
28:28 OK? And again, these are be close by approximal, they're gonna
28:31 upstream, we say farther away. many of these are called enhancer
28:36 Now, if you look at the gene itself, so here's the
28:40 . OK. And under, and see the sequence of exons and
28:47 So that's in a gene, we the areas uh and you carry
28:51 So again, this is all new everything you're seeing here in blue.
28:57 is, this is under Eu Caros don't have genes organized this way.
29:02 the Exxon Enron, so exxons are that are expressed. Enrons are not
29:09 . OK? You have elements called poly A sequence. This, this
29:14 to help the transcript get out of nucleus, for example. So a
29:20 we form was called a primary transcript a pre M R N A that
29:25 basically on a copy of the DNA the exon intra sequences. The there's
29:32 of processing that occurs and you carry uh lots of R N A processing
29:38 transcripts. And so that then becomes spliced, we call it. So
29:45 are removed to produce a continuous coding . So, so you have continuous
29:51 sequences together, OK. And that a translatable transcript. So you get
29:56 of the enrons and you splice together axons. OK. Then you have
30:01 add on things like you see there , the five prime cap the poly
30:07 tail. These are elements that are to uh enhance the stabilization of the
30:15 . It facilitates the uh exit of transcript from the nucleus. Or remember
30:20 you carry out seven nuclear where this transcription occurs and those transcripts must exit
30:25 translation occurs outside the nucleus. So remember that with bacteria and
30:32 they don't have a nucleus. transcription and translation occurs virtually at the
30:36 time. OK? But it's, it is separated in eukaryotes, transcription
30:41 the nucleus translation outside the nucleus. um um so lots of so everything
30:49 know uh under this dash line, that you see below, that is
30:55 you carry out. None, none this occurs in procaryotic types.
31:01 Now, there may be some there's evidence that archaea have some elements
31:06 this. OK. But you for most everything, uh most all
31:13 it's has none of this at OK? And so um so if
31:20 look at procaryote structure, so you have the single gene structure but many
31:26 the bacterial genes, Archeo genes are in the operon, right?
31:31 and it, and it produces a efficient system. OK. So here's
31:34 example of uh one of an OK. So you have a
31:39 of course, you have structural The structural genes are those genes that
31:46 the proteins that do some sort of . And your operon are typically the
31:51 genes will all be common in the function. So they may be participating
31:54 that particular metabolic pathway. OK. chapter 10, we're gonna look at
31:58 lactose operon and crypto operon. So lactose operon contains genes that are specific
32:04 metabolizing glutose lactose. Uh crypto operon genes specific for um producing crypto the
32:12 acids. So that's what OPERON There are groups of genes that are
32:16 to a particular metabolic pathway, And it's a very efficient system because
32:21 can then turn them on all at time and you can turn them off
32:25 at one time. So it provides efficient control of metabolic pathways.
32:31 So the structural genes, I'm the promoter courses, any promoter,
32:38 where a a plym race binds, ? So a a plum races look
32:42 promoters because that will orient the plym in front of the gene,
32:48 And that's where you'll want to begin . OK. So it promotes,
32:51 essential for that reason. Um The of structural genes of course leads to
32:59 formation of the transcript M R N , right? So we call a
33:03 message poly, multiple cyst gene, genes that are part of this same
33:10 . So they're all, it's one transcript. OK. Translation that occurs
33:17 produce proteins, right? If this metabolic pathway, these could be
33:22 enzymes A B and C and that be part of a pathway product.
33:27 reactant W catalyzed by enzyme H produce X which then is acted on by
33:36 and catalyzed it to product Y and forth or a typical metabolic pathway.
33:41 . Um Control elements. So the , right, the operator is a
33:48 element, an OPERON. OK. comes right after the promoter and it
33:52 interact with a regulatory protein. So uh so the OPERON itself, as
33:58 define, it contains these elements, promoter, the operator and the structural
34:04 that defines the operon. OK. regulatory element can be, you
34:09 upstream of this. OK. And it's kind of not considered part of
34:14 operon, but of course, can't it. And so here would be
34:17 regulatory gene that doesn't have to be proximity to this. OK? But
34:22 would code for uh a regulatory protein would then interact with the operator,
34:32 ? And then that can serve to transcription. So now you basically have
34:36 , a um regulatory protein bound to operator that's physically blocking the ability of
34:43 ply to transcribe the structural gene. that's a, that's a, that's
34:48 control element. OK. And um will uh and we'll see in chapter
34:56 , how different conditions can arise, will allow the regulatory protein to bind
35:03 the operator and in some cases not bind and you get transcription. So
35:07 uh that's how control occurs here. , the point is that if you
35:12 expression, you're gonna block expression of those structural genes And shut down the
35:18 pathway, right, or turn it if that's necessary, right? So
35:22 conditions can turn it on other conditions turn it off and that's what we'll
35:26 in chapter 10. OK. Um that's the opera structure, the regular
35:35 , right? So simple factors will in, In a second in Chapter
35:43 . OK. So a sigma factor this right? So remember that Ali
35:49 buying to promoters but they do so the action of a sigma factor,
35:53 ? So in in pro a plum , the sigma factor is part of
36:00 structure. And the sigma factor is guides the R N A lyra to
36:04 promoter. So the Sigma factor will will recognize elements of the promoter and
36:10 to it and then that will help it brings the lyra with it to
36:15 , it binds the promoter and then soon as that binding occurs and the
36:18 begins to transcribe the Sigma factor falls and then can participate in another uh
36:26 round. OK. So that's, what super factors do. And so
36:30 factors then can control multiple operon. . So when you have operon that
36:41 similar control elements, right, that recognize the same Sigma factor, for
36:48 , then that's happening because those OPERON part of a larger common metabolism if
36:57 will. So here's an example, um nitrogen Regulon. So there's one
37:05 the nitrogen Regulon, which kind of the global global nitrogen metabolism in the
37:13 . Ok. So think of just element in nitrogen and all the places
37:19 you find in in in the in in the cell, you find it
37:23 course in amino acids, right? the production of proteins, you find
37:27 in uh nucleotides, right? So the for nucleotide synthesis as well.
37:32 assimilation of nitrogen for different sources, ? So there's all different pathways in
37:38 nitrogen is used and you can kind control it through this global Regulon called
37:44 this case called the nitrogen Regulon. so it kind of serves to control
37:50 this example, the metabolism of So it's a very uh important um
37:57 because again, are highly efficient, ? And they're going to tightly control
38:02 functions uh because one thing and, the reason for this, because it's
38:07 something you can really visualize in any of textbook diagram of transcription and
38:13 And that is the tremendous energy input requires OK. It requires lots of
38:20 to to synthesize a transcript, it lots of energy to translate a
38:27 right. So it's not a trivial . So a a bacterium that is
38:33 protein synthesis, it's it's committing to as long as the conditions are right
38:39 it enable it to happen. Gene . That's why gene control is so
38:43 because if it were to waste gene to, to express genes when it
38:49 need to, that's a tremendous loss energy. And so you have to
38:53 that in nature, you know, Akea are competing with each other in
38:58 other life forms, in their particular environments. And they have to be
39:03 for survival purposes and um to be , turning on genes and turning off
39:09 when they don't need turning on when they don't need to is a
39:12 waste of energy and resources. And they're gonna be efficient to, to
39:16 um for their own survival. And , uh um so they have really
39:22 controlled and in different and in different , right, you can control
39:26 of course, within an opera, you can control multiple opera. So
39:30 all these to, you know, a very efficient system. OK?
39:36 So again, Regulon are under the of multiple operon and those operant presumably
39:41 part of a common global network if will, right? In this
39:46 I gave you to control of nitrogen the cell. OK. So the
39:53 , so plasmids, OK, are extra chromosomal elements. Uh They're of
40:03 much smaller than the chromosome on the of you see, they're 2 to
40:07 kilobytes pairs in size. There are that are upwards of 5 50,000,
40:11 100,000. Uh but most aren't, that big? They may contain uh
40:17 few genes, maybe 1 to you know, some something can contain
40:22 a, a particular uh genes have particular pathway, for example, metabolic
40:28 , uh antibiotic resistant genes. Um about plasmin is they, they have
40:34 own or right. So remember the is the ordinary replication? So if
40:41 have an a that that the can replicate on its own,
40:48 So the plasma is not tied to chromosome. So remember when the the
40:53 , the bacterial cell, the pro will begin to replicate when that cell
40:56 going to divide, right. So is not tied to that, the
41:01 can basically copy itself whenever it wants . OK. Although there is some
41:06 over that. Um but uh the is that the plans are independent of
41:12 . Uh hence the term autonomous, . Um As you see there,
41:17 , the this is actually an artificial constructed in the lab. Uh But
41:22 is based on natural lab. The nat naturally all naturally occur. This
41:27 where they came from. We brought into the lab back in the seventies
41:31 they were maybe late sixties when they discovered. And we've since uh construct
41:36 own plasmids nowadays, of course, elements of natural plasmids. Um but
41:42 a, they're a tool in, , in molecular biology as I'm
41:45 you know, but nonetheless, in plan, you see the uh for
41:50 , you see amp a MP and , these are both antibiotic resistance
41:58 Um These terms see PST one hindi , these are restriction enzymes that where
42:05 can cleave the plasmid and insert different into. Uh but for, you
42:12 , for naturally occurring plasmin, these contain, it could contain antibiotic resistance
42:18 that you see there. Uh these what are called um uh R
42:23 R for resistance that you see at bottom F factors. These are the
42:28 of plasmids that are transferable between right? So the F factor can
42:32 a part of an R factor as . So having the F factor in
42:36 plasma makes it able to be We call it being able to be
42:40 from cell to cell. OK. that we'll talk about in Chapter
42:45 right? That's an integral part of , the F Factor. OK.
42:50 Canta bolic plasmids. And so just make a point. So your your
42:53 factors subs that contain antiac resistance or that contain canna bolic pathways in
42:59 These can also contain F factors that them mobile, then it can be
43:03 . OK. Um And so can plasmas can, may contain a a
43:09 . Uh So the the the uh inheriting pathetic can acquire this this potentially
43:16 catalog pathway. OK. So plasmids are not um the cells that have
43:24 , of course, can't provide a under certain conditions, right? But
43:28 plasmas are not critical for survival unless course, they're faced with the condition
43:36 the plasma, the gene product of plasma is one that enables survival.
43:42 . So, in in the presence antibiotic, if that antibiotic is
43:46 then of course, those cells with plasma will survive. OK. And
43:50 that antibiotic is not present, you , it, it it does not
43:53 give it any benefit really. But , um uh if we look at
43:59 replication of plasmids, OK. We're with the way the chromosome replicates and
44:05 replicate this as a way as right? The bidirectional replication mechanism,
44:09 , where you have the the uh separation at the origin um and then
44:16 uh each strand, right, the bidirectional model, semiconservative replication and then
44:24 formation of two identical guinea molecules. . The rolling circle replication,
44:30 That was is unique to plasmids. . And uh that we typically see
44:39 prior to cells that are going to , right? When that plasmid replicates
44:44 is given to another cell. And so what happens is a process
44:52 the nick formation. OK. So have a plasmid and we we break
44:58 covalent bond in that um sugar phosphate of the DNA. OK. And
45:06 doing so, we expose a three hydroxyl answer. So remember your ira
45:15 , right? So the DNA plum DNA. So D A plum
45:19 it functions by being able to extend a three prime hydroxyl. OK.
45:26 it can add nucleated types to that . And so what happens in arow
45:31 replication? A nick is made to that covalent bond that exposed through P
45:37 and is then the APR can buy that and begin to copy. So
45:43 copy right. And you see that complementary strand is being displaced, right
45:50 shown right there, right? So being displaced as new DNA is being
46:00 to that template. And so as continues around the strand, then that
46:07 is displaced as you see here. . And what happens in conjugation is
46:14 would have two cells, one here one here. OK. And then
46:23 other DNA as is being displaced would funneled into the cell it's mating
46:30 right? And now that cell acquires copy of that plasma. OK?
46:34 that initially single strano plasmid that was displaced is copied as well to make
46:42 double strand as you see there. . So that's rolling circle replication.
46:47 gonna see that as we talk about , as we get into chapter nine
46:50 look at conjugation how 22 cells come in kind of a pseudo mating
46:58 Although there's not, there's not sexual by any means, it's just a
47:03 of the type of um replication and a type of of gene transfer
47:09 So two cells come together and then cell will carry a royal circle application
47:13 was plasmid, then displaced strand is into the cell it's mating with and
47:20 cell now received a copy of that . So that's so we'll see that
47:24 , the tip. So again, circle application typically is what will precede
47:31 conjugation and it occurs with the plasmid it's being copied. OK? Um
47:39 inherited the plasmids. So of there's selective pressure, right? Um
47:46 a take your back here for a , your um plans, right?
47:55 cell containing this plasmid, right? we have a resistance. Amici R
48:05 is for resistance and tet tetracycline OK. So a a cell will
48:15 onto that plasmin for sure if it's selective pressure. If tetracycline or ampicillin
48:23 present, those cells will survive. so that's selective pressure to keep that
48:28 . OK. So that's what we by selective pressure. OK? If
48:31 contained in a can a a metabolic genes or metabolic pathway, if
48:36 if that particular uh metabolic compound were , then that would uh lead to
48:43 pressure to maintain that plasma. So it's a, that's what we,
48:48 mean by that. And if you uh you know cells that we do
48:54 manipulations with using plasmids and plasma generally have an antibiotic in the resistance gene
49:00 them. And so we, we sure the cells hold not the plasmid
49:04 , by growing them in the presence , of the antibiotic, right.
49:08 applying that selective pressure. OK. so uh and so there's also among
49:16 , there's this feature of low copy high copy number. OK. So
49:24 low copy number plasmids uh maybe have or two copies in the cell at
49:29 . OK. And whether their higher copy has to do with the,
49:33 type of origin and replication they have um uh it, it's just an
49:41 feature of the type of plasma. is OK, whether it's low or
49:44 copy number, he had copy number , for example, something like
49:51 When the cell divides the, you the the the likelihood is very high
49:58 when the cell divides, for like so that each daughter's cell is
50:04 to receive at least one plasmid, ? It was a low copy number
50:12 . Well then that's a little OK? Because even just by random
50:18 , maybe the cell does not receive copy of the plasm because there's only
50:21 present, right. So a mechanism was discovered that can ensure the plasmid
50:28 is passed on to the next Once replication is complete is through these
50:34 are called par proteins. Par is for partition. So partition proteins,
50:39 these are kind of like acting like , OK. That polymerize and kind
50:47 help guide bind to the plasmid and it's copied into the copy of the
50:54 and kind of guide it to each of the cell, each pole of
50:58 cell to ensure that it uh when cell divides, each half of the
51:02 gets a copy of that. We can see it better close up
51:07 . Uh There you see it Let me just go to the detailed
51:10 . So here is um and, it's a energy requiring process. Um
51:16 so, uh and so here you a plasmid and the plasmin copy and
51:23 you see these par proteins part R bind to the plasmid and then to
51:28 are the par m which kind of it kind of acts like a pseudo
51:35 mitotic spindle if you will, it's a mitotic spindle, but it kind
51:38 acts that way, right? So , we're binding these uh par proteins
51:44 then polymerizing and then they bind, join up like, so like you
51:48 here in this Number three Graphic, . And the uh at that point
52:01 as the um filaments elongate, each plasm copy is pushed to opposite
52:06 in the cell. And then that certainly ensure that um the the binding
52:12 the pattern to the pole triggers and breakdown of the filaments. And so
52:18 this, I'm guessing precedes the uh division. And so now the soul
52:24 divide and then that ensures that each gets a copy of the, of
52:30 plasma. So for low copy number , which can be, you
52:34 at one copy per cell. This a mechanism that can kind of ensure
52:37 when it replicates, the daughter cell a copy of the password.
52:44 So, um as we look at chapter eight, we're just gonna look
52:53 the, the, the a, plume and then a little bit about
52:57 and binding sites. So, the plum hollow enzyme has multiple subunits,
53:03 two alpha and two beta subunits. that's typically where this is where
53:07 the uh replication or the transcription of D A and the R N A
53:14 . Um The Sigma factor, as mentioned previously, this is involved in
53:20 recognition and a sigma factor is is a um uh portion of the
53:25 that's not permanent. So, while , the, the two alpha and
53:30 beta sapient are part of the core , the um Sigma factor is one
53:37 binds and unbinds from the core But when they're all together, when
53:41 have the, the two alpha and be uns and the sigma factor
53:45 that's what we call the hollow enzyme the whole enzyme, right? But
53:50 sigma factor, once it, it the plum race to the promoter,
53:54 it can then release and then go to bind another core plym race to
54:00 promoter. OK. So kind of this kind of fashion here. So
54:05 is the Sigma factor looking for? . Well, it recognizes promoter sequences
54:10 the position is -35 -10, which very common in bacteria. OK.
54:17 so you see the uh hollow enzyme is a sigma factor that's scanning the
54:23 B N A looking for this minus minus 10 and motor sequences from check
54:32 once found, then the polymerase unbinds and the core pra initiates um strand
54:42 and transcription. OK. So um when we look at uh a closer
54:51 at the sequences, factors recognize the -35 is a consensus sequence.
54:58 right. So looking at multiple promoter of various bacterial genes, we see
55:03 this is a very common uh right? This consensus sequence is minus
55:10 minus 10 region, right? They to be rich in uh T A
55:20 , thymine adding because remember the thymine is two hydrogen bonds, the quiNINE
55:28 G C is three. So two bonds makes it easier en energy wise
55:33 pull those strands apart and begin So typically they're, they're thymine rich
55:39 for that reason, either thus energy help pull apart to begin transcription.
55:44 The sigma uh 70 factor is very . Most of your bacterial genes utilize
55:50 Sigma factor. But there are And so here you see the consensus
55:55 of several uh promoters, what we are called strong promoters. Um You
56:02 manipulate sequences within the -35-10 to either or increase activity by substituting nucleotides,
56:13 see they're down, up, down down um change means a decrease in
56:19 . The less transcription, up up uh allows for an increase in transcription
56:27 . OK. And so we talk strength, right? Strong versus
56:31 It all relates to the level of , right? So you have strong
56:35 and your weak promoters, right? weak promoters aren't necessarily bad. They're
56:39 weak that that's, that's the way , they, they function is to
56:43 to have less expression. OK. you have to remember a lot of
56:48 , your metabolic pathways are all integrated the products of one are the reactors
56:52 another and so forth. And so will be changes in type levels of
56:57 to keep everything in balance. Um But certainly, you know,
57:02 a biotech technical standpoint, biotechnology you know, when one wants to
57:09 if one has a particular protein that's commercial interest and you want to,
57:14 want the bacterium producing it to over it. Um You, one of
57:20 things you could do is tweak the for the gene producing that protein to
57:25 it to express more that can lead other problems. But you know,
57:29 , it's a way to begin to overproduce the protein. OK?
57:34 any case, so weak versus uh versus weak expression, OK. Relates
57:40 high versus low expression. All it all has to do with the
57:44 of the pliers to the promoter. . So tight binding means lots of
57:51 . Generally weak binding leads to less . OK? And we, we
57:58 about basal level of basal levels of are typically weak, but that gets
58:04 by involving different regulatory elements, Different regulatory proteins can come in to
58:10 levels of expression. OK. So all a part of, of controlling
58:16 uh is the manipulation of promoter OK. So, uh aside from
58:23 Sigma 70 which is one of the common ones, uh you can have
58:28 like sigma 32 heat shock and new . These are often see stress
58:34 not just heat but other types of . These are produced to, to
58:39 to, to turn on certain genes in dealing with that stress. Um
58:45 are Matilda chemotaxis, specific sigma both of these have minus 35 minus
58:53 regions but, but recognize slightly different as you see on the right
58:59 the uh there are several factors involved this uh stationary phase. Um The
59:06 P O N is one in nitrogen and we talked earlier about the example
59:11 Regulon, right. So in, this nitrogen Regulon, then those operon
59:17 in that will have promoters that recognize called a minus 24 minus 12
59:23 So that's specific for nitrogen metabolism uh . Uh So that, that's the
59:30 the Sigma 54 fits that category. just to show you that there's some
59:35 . Uh but the most common for genes is a sigma 70. But
59:38 , we have these differences. So finally, then is the kind
59:44 bring it all together and uh the we talked about in this module.
59:50 the here again is the opera structure you see, you know, keeping
59:55 relatively simple here, we have uh course, our promoter. OK.
60:02 uh minus 35 minus 10 sequence, is common for most bacterial promoters.
60:07 there are variations uh remember the sense the coding strand and the template strand
60:12 talked about that. Uh Here's our structural genes A and B.
60:18 Um The uh expression of that will a single message containing the information for
60:26 genes. So the term polycystic, . So here's uh a gene A
60:33 B uh in that transcript, uh that each has its own uh start
60:41 codon, right, start stop for individual gene. That's part of that
60:47 . OK. Then that will produce Through translation, right? So we
60:53 the ribosome, right? The 70 ribosome is what we find in
60:58 OK. And that's a combination of these small Uh 30 s subunit and
61:05 50 s subunit that come together to the functioning ribosome that translates OK.
61:13 there's a Robertson binding site, That's shown there. So that's what
61:17 call the Shine Del Garno sequence named those that discovered it. And so
61:22 binds to the 16 SRN A, ? We talked about 16 SRN A
61:26 a different context uh in terms of . Uh but it's functionally, it
61:31 serves to bind to the ra on site. OK. And begin the
61:38 of translation. OK. So, so then translation brings about the the
61:51 of the two proteins. This would A and be OK. So,
62:01 um so just uh so think about elements needed for each process,
62:06 For, for transcription, you need promoter. OK. Um The uh
62:15 factor is involved in transcription, binding the promoter. Uh most your Vetri
62:21 at -35 -10 sequence. Um We're operon here. So, right,
62:28 we're gonna have uh multiple genes under control of one promoter. Uh the
62:34 process, right involves uh uh or sorry, transcription then of the,
62:40 I remember the transcription of the which copying the template strand, right?
62:45 then that M R N A is the the coding strand, it contains
62:49 coding information. OK. Then to that uh transcript, you know,
62:56 transfer R N A S, the binding to a ribosome binding site.
63:03 . And initiating transcript a translation, . And then the production of the
63:11 peptides from that transcript. OK. just making sure you got the,
63:17 components that the, the, the of what happens at each stage and
63:20 involved. OK. And knowing what where, right. So similar factors
63:26 involved around own binding. It's, uh no sequence, right? That
63:32 the is uh so just make sure have those differentiated. Here's what be
63:38 in transcription, here's what blocks in , uh et cetera. OK.
63:45 uh and then remember the poly rhizome , which I talked about in chapter
63:50 , so that there is no nucleus the pro right. So transcription and
63:56 can occur virtually at the same OK. As, as a as
64:03 uh uh Rome is transcribing a Then as soon as that ribosome binding
64:13 of the transcript appears ribosomes will begin and translating. So you're gonna have
64:19 ribosomes on a transcript that are transcribing the the polyribosome or polysome formation
64:27 So, so that's then summarizes chapter and 8 then Or concludes chapter 7
64:34 . And so again, it just basics of um The operon structure.
64:41 Regulon are OK, plasmids, some in the chromosome together can constitute the
64:47 of a bacterium. Uh if they plasmas, of course, uh in
64:52 eight, you know, the the of the sigma factor right is in
64:57 , the role of the of the shine dear sequence is his own
65:03 OK. For, for translation uh , we didn't talk about termination of
65:09 . So just please Ignore that. so uh so in the next
65:17 then we'll continue on with Chapter which will be um looking at different
65:25 of gene transfer, right. bacteria can of course divide by binary
65:30 and other cells inherit gene that But that's one mode. The other
65:34 is uh cells can acquire DNA from environment. They can acquire DNA by
65:44 with other cells in the population um through viruses, they can acquire New
65:51 . So look for that in Chapter . OK. Thanks
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