which of the following conditions increase the expression of lacz from the lac operon
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get which of the following conditions increase the expression of lacz from the lac operon from EN Bilgi.
Lac Operon
Lac Operon
The lac operon exhibits both negative control and positive control.
From: Elsevier's Integrated Review Biochemistry (Second Edition), 2012
Related terms:
LactosePlasmidLac RepressorPhosphoproteinRepressorNested GeneMutationEscherichia coli
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lac Operon
L. Swint-Kruse, K.S. Matthews, in Encyclopedia of Biological Chemistry (Second Edition), 2013
Multiple Sites/Multiple Targets – DNA Looping
In addition to the primary operator, LacO, shown in Figure 1, two ‘pseudo-operator’ sequences are present within the lac operon sequence and contribute to repression. The DNA sequences of the pseudo-operators are very similar, but not identical, to LacO and are bound by LacI more weakly. The presence of two DNA-binding sites in LacI protein tetramer suggested a mechanism by which pseudo-operators could enhance repression – one LacI tetramer could bind two separate operators and generate a looped DNA structure. Experimental evidence for DNA looping has been obtained from a variety of laboratories. Recent evidence indicates that the angle between the two dimers must open for loop formation to occur, as illustrated in Figure 4. These looped structures are highly stabilized, accounting for the significant repression of lacZYA expression observed in bacterial cells. Indeed, DNA containing multiple operator sequences and with the supercoiling density characteristic of E. coli exhibits a half-life for the complex that exceeds 2 days. However, even these looped complexes respond rapidly (in less than 30 s) to the presence of inducer sugars, allowing quick adaptation to an external lactose source that may be transient.
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Figure 4. Looped DNA structure. The teal blue curved line depicts the lac operon DNA (with shading to indicate nearness to observer), which contains three possible LacI-binding sites (two of which, O1 and O2, are shown bound to LacI). The pseudo-operator sequence O2 is located within the lacZ gene, and the primary operator sequence O1 overlaps the promoter sequence for the lacZYA metabolic genes (Figure 1). Tetrameric LacI is shown at the bottom of the figure as simultaneously interacting with O1 and O2. This structure loops the DNA and generates a complex with very high stability. Note that the dimers within a LacI tetramer separate and adopt a larger angle between them when looping between O1 and O2 than in the absence of looping (i.e., the structure shown in Figure 3(a)). The need for flexibility between the dimers for looping to occur is supported by experimental evidence.
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lac Operon
Liskin Swint-Kruse, Kathleen S. Matthews, in Encyclopedia of Biological Chemistry, 2004
LacI Function
The role of LacI is to inhibit mRNA production for proteins encoded by the lac operon. Transcription is not completely eliminated, but lacZYA mRNA is transcribed only at very low levels. This function is accomplished by specific binding of LacI protein to the lac operator DNA sequence to inhibit transcription via a variety of mechanisms. Since the lac operator (LacO) overlaps the promoter, LacI binding directly competes with RNA polymerase for binding this region. LacI can also impede transcription initiation and/or block elongation of mRNA. The LacI·LacO association and consequent transcription repression occur when no lactose is available to serve as the substrate of the lac metabolic proteins.
When lactose is available as a carbon source, the low levels of metabolic enzymes allow a small amount of this sugar to be transported into the bacterium by LacY. Next, residual LacZ metabolizes lactose to glucose and galactose, which produces energy for the bacterium. Notably, this catalytic process also generates low levels of allolactose (a rearrangement of the β-1,4 linkage between glucose and galactose to a β-1,6 linkage; Figure 2). The by-product allolactose binds to LacI and elicits a conformational change in the protein that results in release of the operator DNA sequence (induction). Consequently, RNA polymerase is freed to generate numerous copies of mRNA encoding the lac enzymes. When translated into proteins, these enzymes allow the bacterium to transport and metabolize large quantities of lactose as its carbon energy source, taking advantage of environmental opportunity. One result of the studies by Jacob and Monod was the discovery that a variety of non-natural galactoside sugars (e.g., IPTG; Figure 2) can induce LacI and relieve transcription repression of lacZYA.
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Figure 2. Chemical structures for lactose, allolactose (the natural inducer), the sugar components of lactose (glucose and galactose), the gratuitous inducer IPTG, and cAMP.
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RNA Transcription and Control of Gene Expression
John W. Pelley, in Elsevier's Integrated Review Biochemistry (Second Edition), 2012
Regulation of the Lac Operon
The lack of a nuclear membrane in prokaryotes gives ribosomes direct access to mRNA transcripts, allowing their immediate translation into polypeptides. This makes transcription the rate-limiting step in prokaryotic gene expression and, therefore, a major point of regulation. The classic example of prokaryotic gene regulation is that of the lac operon. This operon is a genetic unit that produces the enzymes necessary for the digestion of lactose (Fig. 16-13).
Source : www.sciencedirect.com
12.1: The lac Operon
Early insights into mechanisms of transcriptional regulation came from studies of E. coli by researchers Francois Jacob & Jacques Monod. In E. coli, and many other bacteria, genes encoding …
12.1: The lac Operon
Last updated Apr 9, 2022
12: Regulation of Gene Expression
12.2: The Use of Mutants to Study the lac Operon
Todd Nickle and Isabelle Barrette-Ng
Mount Royal University & University of Calgary
Early insights into mechanisms of transcriptional regulation came from studies of E. coli by researchers Francois Jacob & Jacques Monod. In E. coli, and many other bacteria, genes encoding several different proteins may be located on a single transcription unit called an operon. The genes in an operon share the same transcriptional regulation, but are translated individually. Eukaryotes generally do not group genes together as operons (exception is C. elegans and a few other species).
Basic lac Operon structure
E. coli encounters many different sugars in its environment. These sugars, such as lactose and glucose, require different enzymes for their metabolism. Three of the enzymes for lactose metabolism are grouped in the lac operon: lacZ, lacY, and lacA (Figure
12.1.1 12.1.1
). LacZ encodes an enzyme called β-galactosidase, which digests lactose into its two constituent sugars: glucose and galactose. lacY is a permease that helps to transfer lactose into the cell. Finally, lacA is a trans-acetylase; the relevance of which in lactose metabolism is not entirely clear. Transcription of the lac operon normally occurs only when lactose is available for it to digest. Presumably, this avoids wasting energy in the synthesis of enzymes for which no substrate is present. A single mRNA transcript includes all three enzyme-coding sequences and is called polycistronic. A cistron is equivalent to a gene.
Figure 12.1.1 12.1.1
: Diagram of a segment of an E. coli chromosome containing the lac operon, as well as the lacI coding region. The various genes and cis-elements are not drawn to scale. (Originall-Deyholos-CC:AN)
cis- and transRegulators
In addition to the three protein-coding genes, the lac operon contains short DNA sequences that do not encode proteins, but are instead binding sites for proteins involved in transcriptional regulation of the operon. In the lac operon, these sequences are called P (promoter), O (operator), and CBS (CAP-binding site). Collectively, sequence elements such as these are called cis-elements because they must be located on the same piece of DNA as the genes they regulate. On the other hand, the proteins that bind to these cis-elements are called trans-regulators because (as diffusible molecules) they do not necessarily need to be encoded on the same piece of DNA as the genes they regulate.
lacI is an allosterically regulated repressor
One of the major trans-regulators of the lac operon is encoded by lacI. Four identical molecules of lacI proteins assemble together to form a homotetramer called a repressor (Figure
12.1.2 12.1.2
). This repressor binds to two operator sequences adjacent to the promoter of the lac operon. Binding of the repressor prevents RNA polymerase from binding to the promoter (Figure
12.1.3 12.1.3
). Therefore, the operon will not be transcribed when the operator is occupied by a repressor.
Figure 12.1.2 12.1.2
: Structure of lacI homotetramer bound to DNA (Origianl-Deyholos-CC:AN)
Besides its ability to bind to specific DNA sequences at the operator, another important property of the lacI protein is its ability to bind to lactose. When lactose is bound to lacI, the shape of the protein changes in a way that prevents it from binding to the operator. Therefore, in the presence of lactose, RNA polymerase is able to bind to the promoter and transcribe the lac operon, leading to a moderate level of expression of the lacZ, lacY, and lacA genes. Proteins such as lacI that change their shape and functional properties after binding to a ligand are said to be regulated through an allosteric mechanism. The role of lacI in regulating the lac operon is summarized in Figure
12.1.4 12.1.4 .
Figure 12.1.3 12.1.3
: When the concentration of lactose [Lac] is low, lacI tetramers bind to operator sequences (O), thereby blocking binding of RNApol (green) to the promoter (P). Alternatively, when [Lac] is high, lactose binds to lacI, preventing the repressor from binding to O, and allowing transcription by RNApol. (Origianl-Deyholos-CC:AN)
CAP is an allosteric activator of the lac operon
A second aspect of lac operon regulation is conferred by a trans-factor called cAMP binding protein (CAP, Figure
12.1.4 12.1.4
). CAP is another example of an allosterically regulated trans-factor. Only when the CAP protein is bound to cAMP can another part of the protein bind to a specific cis-element within the lac promoter called the CAP binding sequence (CBS). CBS is located very close to the promoter (P). When CAP is bound to at CBS, RNA polymerase is better able to bind to the promoter and initiate transcription. Thus, the presence of cAMP ultimately leads to a further increase in lac operon transcription.
The lac operon (article)
Regulation of genes for lactose utilization. lac repressor, catabolite activator protein, and cAMP.
Regulation of gene expression and cell specialization
The lac operon
Regulation of genes for lactose utilization. lac repressor, catabolite activator protein, and cAMP.
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Key points:
The lac operon of E. coli contains genes involved in lactose metabolism. It's expressed only when lactose is present and glucose is absent.
Two regulators turn the operon "on" and "off" in response to lactose and glucose levels: the lac repressor and catabolite activator protein (CAP).
The lac repressor acts as a lactose sensor. It normally blocks transcription of the operon, but stops acting as a repressor when lactose is present. The lac repressor senses lactose indirectly, through its isomer allolactose.
Catabolite activator protein (CAP) acts as a glucose sensor. It activates transcription of the operon, but only when glucose levels are low. CAP senses glucose indirectly, through the "hunger signal" molecule cAMP.Introduction
Lactose: it's what's for dinner! While that may not sound delicious to us (lactose is the main sugar in milk, and you probably don't want to eat it plain), lactose can be an excellent meal for E. coli bacteria. However, they'll only gobble up lactose when other, better sugars – like glucose – are unavailable.
With that for context, what exactly is the lac operon? The lac operon is an operon, or group of genes with a single promoter (transcribed as a single mRNA). The genes in the operon encode proteins that allow the bacteria to use lactose as an energy source.
What makes the lac operon turn on?
E. coli bacteria can break down lactose, but it's not their favorite fuel. If glucose is around, they would much rather use that. Glucose requires fewer steps and less energy to break down than lactose. However, if lactose is the only sugar available, the E. coli will go right ahead and use it as an energy source.
To use lactose, the bacteria must express the lac operon genes, which encode key enzymes for lactose uptake and metabolism. To be as efficient as possible, E. coli should express the lac operon only when two conditions are met:
Lactose is available, and
Glucose is not available
How are levels of lactose and glucose detected, and how how do changes in levels affect lac operon transcription? Two regulatory proteins are involved:
One, the lac repressor, acts as a lactose sensor.
The other, catabolite activator protein (CAP), acts as a glucose sensor.
These proteins bind to the DNA of the lac operon and regulate its transcription based on lactose and glucose levels. Let's take a look at how this works.
Structure of the lac operon
The lac operon contains three genes: lacZ, lacY, and lacA. These genes are transcribed as a single mRNA, under control of one promoter.
Genes in the lac operon specify proteins that help the cell utilize lactose. lacZ encodes an enzyme that splits lactose into monosaccharides (single-unit sugars) that can be fed into glycolysis. Similarly, lacY encodes a membrane-embedded transporter that helps bring lactose into the cell. [More details]
In addition to the three genes, the lac operon also contains a number of regulatory DNA sequences. These are regions of DNA to which particular regulatory proteins can bind, controlling transcription of the operon.
Structure of the lac operon. The DNA of the lac operon contains (in order from left to right): CAP binding site, promoter (RNA polymerase binding site), operator (which overlaps with promoter), lacZ gene, lacY gene, and lacA gene. The activator protein CAP, when bound to a molecule called cAMP (discussed later), binds to the CAP binding site and promotes RNA polymerase binding to the promoter. The lac repressor protein binds to the operator and blocks RNA polymerase from binding to the promoter and transcribing the operon.
_Image modified from "Prokaryotic gene regulation: Figure 3," by OpenStax College, Biology (CC BY 4.0)._
The promoter is the binding site for RNA polymerase, the enzyme that performs transcription.
The operator is a negative regulatory site bound by the lac repressor protein. The operator overlaps with the promoter, and when the lac repressor is bound, RNA polymerase cannot bind to the promoter and start transcription.
The CAP binding site is a positive regulatory site that is bound by catabolite activator protein (CAP). When CAP is bound to this site, it promotes transcription by helping RNA polymerase bind to the promoter.
Let's take a closer look at the lac repressor and CAP and their roles in regulation of the lac operon.
The lac repressor
The lac repressor is a protein that represses (inhibits) transcription of the lac operon. It does this by binding to the operator, which partially overlaps with the promoter. When bound, the lac repressor gets in RNA polymerase's way and keeps it from transcribing the operon. [Where does the lac repressor come from?]
When lactose is not available, the lac repressor binds tightly to the operator, preventing transcription by RNA polymerase. However, when lactose is present, the lac repressor loses its ability to bind DNA. It floats off the operator, clearing the way for RNA polymerase to transcribe the operon.
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