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The lac operon

AP.BIO:
IST‑2 (EU)
,
IST‑2.A (LO)
,
IST‑2.A.1 (EK)
,
IST‑2.A.3 (EK)
,
IST‑2.B (LO)
,
IST‑2.B.1 (EK)
,
IST‑2.C (LO)
,
IST‑2.C.1 (EK)
,
IST‑2.C.2 (EK)
Regulation of genes for lactose utilization. lac repressor, catabolite activator protein, and cAMP.

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.
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.
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.
Upper panel: No lactose. When lactose is absent, the lac repressor binds tightly to the operator. It gets in RNA polymerase's way, preventing transcription.
Lower panel: With lactose. Allolactose (rearranged lactose) binds to the lac repressor and makes it let go of the operator. RNA polymerase can now transcribe the operon.
_Image modified from "Prokaryotic gene regulation: Figure 3," by OpenStax College, Biology (CC BY 4.0)._
This change in the lac repressor is caused by the small molecule allolactose, an isomer (rearranged version) of lactose. When lactose is available, some molecules will be converted to allolactose inside the cell. Allolactose binds to the lac repressor and makes it change shape so it can no longer bind DNA.
Allolactose is an example of an inducer, a small molecule that triggers expression of a gene or operon. The lac operon is considered an inducible operon because it is usually turned off (repressed), but can be turned on in the presence of the inducer allolactose.

Catabolite activator protein (CAP)

When lactose is present, the lac repressor loses its DNA-binding ability. This clears the way for RNA polymerase to bind to the promoter and transcribe the lac operon. That sounds like the end of the story, right?
Well...not quite. As it turns out, RNA polymerase alone does not bind very well to the lac operon promoter. It might make a few transcripts, but it won't do much more unless it gets extra help from catabolite activator protein (CAP). CAP binds to a region of DNA just before the lac operon promoter and helps RNA polymerase attach to the promoter, driving high levels of transcription.
Upper panel: Low glucose. When glucose levels are low, cAMP is produced. The cAMP attaches to CAP, allowing it to bind DNA. CAP helps RNA polymerase bind to the promoter, resulting in high levels of transcription.
Lower panel: High glucose. When glucose levels are high, no cAMP is made. CAP cannot bind DNA without cAMP, so transcription occurs only at a low level.
_Image modified from "Prokaryotic gene regulation: Figure 3," by OpenStax College, Biology (CC BY 4.0)._
CAP isn't always active (able to bind DNA). Instead, it's regulated by a small molecule called cyclic AMP (cAMP). cAMP is a "hunger signal" made by E. coli when glucose levels are low. cAMP binds to CAP, changing its shape and making it able to bind DNA and promote transcription. Without cAMP, CAP cannot bind DNA and is inactive.
CAP is only active when glucose levels are low (cAMP levels are high). Thus, the lac operon can only be transcribed at high levels when glucose is absent. This strategy ensures that bacteria only turn on the lac operon and start using lactose after they have used up all of the preferred energy source (glucose).

So, when does the lac operon really turn on?

The lac operon will be expressed at high levels if two conditions are met:
  • Glucose must be unavailable: When glucose is unavailable, cAMP binds to CAP, making CAP able to bind DNA. Bound CAP helps RNA polymerase attach to the lac operon promoter.
  • Lactose must be available: If lactose is available, the lac repressor will be released from the operator (by binding of allolactose). This allows RNA polymerase to move forward on the DNA and transcribe the operon.
These two events in combination – the binding of the activator and the release of the repressor – allow RNA polymerase to bind strongly to the promoter and give it a clear path for transcription. They lead to strong transcription of the lac operon and production of enzymes needed for lactose utilization.

Putting it all together

Now that we’ve seen all the moving parts of the lac operon, let’s put what we’ve learned together to see how the operon reacts to a variety of different conditions (presence or absence of glucose and lactose).
  • Glucose present, lactose absent: No transcription of the lac operon occurs. That's because the lac repressor remains bound to the operator and prevents transcription by RNA polymerase. Also, cAMP levels are low because glucose levels are high, so CAP is inactive and cannot bind DNA.
    Glucose present, lactose absent: No transcription of the lac operon occurs. That's because the lac repressor remains bound to the operator and prevents transcription by RNA polymerase. Also, cAMP levels are low because glucose levels are high, so CAP is inactive and cannot bind DNA.
    _Image modified from "Prokaryotic gene regulation: Figure 3," by OpenStax College, Biology (CC BY 4.0)._
  • Glucose present, lactose present: Low-level transcription of the lac operon occurs. The lac repressor is released from the operator because the inducer (allolactose) is present. cAMP levels, however, are low because glucose is present. Thus, CAP remains inactive and cannot bind to DNA, so transcription only occurs at a low, leaky level.
    Glucose present, lactose present: Low-level transcription of the lac operon occurs. The lac repressor is released from the operator because the inducer (allolactose) is present. cAMP levels, however, are low because glucose is present. Thus, CAP remains inactive and cannot bind to DNA, so transcription only occurs at a low, leaky level.
    _Image modified from "Prokaryotic gene regulation: Figure 3," by OpenStax College, Biology (CC BY 4.0)._
  • Glucose absent, lactose absent: No transcription of the lac operon occurs. cAMP levels are high because glucose levels are low, so CAP is active and will be bound to the DNA. However, the lac repressor will also be bound to the operator (due to the absence of allolactose), acting as a roadblock to RNA polymerase and preventing transcription.
    Glucose absent, lactose absent: No transcription of the lac operon occurs. cAMP levels are high because glucose levels are low, so CAP is active and will be bound to the DNA. However, the lac repressor will also be bound to the operator (due to the absence of allolactose), acting as a roadblock to RNA polymerase and preventing transcription.
    _Image modified from "Prokaryotic gene regulation: Figure 3," by OpenStax College, Biology (CC BY 4.0)._
  • Glucose absent, lactose present: Strong transcription of the lac operon occurs. The lac repressor is released from the operator because the inducer (allolactose) is present. cAMP levels are high because glucose is absent, so CAP is active and bound to the DNA. CAP helps RNA polymerase bind to the promoter, permitting high levels of transcription.
    Glucose absent, lactose present: Strong transcription of the lac operon occurs. The lac repressor is released from the operator because the inducer (allolactose) is present. cAMP levels are high because glucose is absent, so CAP is active and bound to the DNA. CAP helps RNA polymerase bind to the promoter, permitting high levels of transcription.
    _Image modified from "Prokaryotic gene regulation: Figure 3," by OpenStax College, Biology (CC BY 4.0)._

Summary of lac operon responses

GlucoseLactoseCAP bindsRepressor bindsLevel of transcription
+--+No transcription
++--Low-level transcription
--++No transcription
-++-Strong transcription

Explore outside of Khan Academy

Do you want to learn more about the lac operon? Check out this scrollable interactive from LabXchange.
LabXchange is a free online science education platform created at Harvard’s Faculty of Arts and Sciences and supported by the Amgen Foundation.

Want to join the conversation?

  • blobby green style avatar for user 😊
    Lactose enter into cell with Help of permease....but permease enzyme is produced by lactose? Which one is first. Lac or permease?
    (8 votes)
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    • orange juice squid orange style avatar for user alannah.king13
      Although when the repressor is bound (Or when CAP is unbound) transcription becomes incredibly difficult, it still occurs but just very, very inefficiently. So there will be tiny amounts of permease produced normally through these rare chance events, which can "kick start" the process if there happens to be lactose outside the cell :)
      (18 votes)
  • mr pink red style avatar for user Revan Rangotis
    If the expression of the lac operon is induced by the isomer of lactose, allolactose, and beta-galactosidase, the protein product of this operon, is the enzyme responsible for lactose isomerisation, where does the initial allolactose come from?

    Is it a natural mechanism, as in, in happens spontaneously, or is some of the beta-galactosidase present in the cell anyway?
    (4 votes)
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    • female robot grace style avatar for user tyersome
      Very good question!

      To understand this it helps to keep in mind that very few things in biology are 100% — this sloppiness or leakiness turns out be absolutely critical for many biological processes.

      In the case of the lac operon, there is a low level of expression even when it is shut-off by the lac repressor. This means a small amount of the ß-galactosidase (encoded by lacZ ) is always present.
      (The low level expression of the symporter encoded by lacY (ß-galactoside permease) is also required to allow lactose into the cell.)

      There is a very extensive Wikipedia article on this:
      https://en.wikipedia.org/wiki/Lac_operon
      (5 votes)
  • blobby green style avatar for user Christina Lynn
    how are E. coli able to use up all of the glucose present before turning to lactose?
    (2 votes)
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  • piceratops ultimate style avatar for user Bailan
    Is lac operon only related to lactose metabolism in E.coli?
    Does humans need any such operon to metabolise lactose??
    (1 vote)
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  • piceratops sapling style avatar for user http://facebookid.khanacademy.org/1476580007
    Why is lac operon so important in modern molecular biology?
    (1 vote)
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    • blobby green style avatar for user amconnel99
      Great question. The reason I have found that the lac operon is so important, is that it is the most study operon and has become the most classic example of how an operon works. My biology teacher for AP said that is the most common example. Hope that helps!
      (5 votes)
  • blobby green style avatar for user isabellewild01
    what happens if the repressor is is mutated and cannot bind to the operator. How would this affect transcription when both glucose and lactose are present
    (3 votes)
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  • blobby green style avatar for user toadere17
    If genes in an operon are transcribed together how does translation occur? Is there a mechanism in place that separates the different proteins or a long chain of aa is made and the different proteins are then further separated?
    (2 votes)
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    • female robot grace style avatar for user tyersome
      Good question!


      Even though all the genes in an operon are transcribed together, they are not translated together.

      The ORF (open reading frame) for each gene is terminated by a stop codon and there often are untranslated sequences between the ORFs for each gene.

      At the start of each gene there will be a RBS (ribosome binding site).

      When the mRNA is being translated, the RBSs for each gene will independently§ bind ribosomes and thus each gene makes a separate protein.


      This figure may help make things clearer:
      https://en.wikipedia.org/wiki/Operon#Overview


      Does that help?


      §Note: The RBS for a "downstream" gene may bind a separate ribosome or it may bind a ribosome that just came off of the "upstream" gene.
      (1 vote)
  • blobby green style avatar for user Noaamir17
    does the suppressor regulate the cap-Camp complex?
    (2 votes)
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    • winston baby style avatar for user Ivana - Science trainee
      CAP binds the CAP binding site of the lac promoter to carry out negative control of operon gene transcription, whereas cAMP blocks the CAP binding site and thereby allows fine-tuning of the system.

      Glucose control is accomplished because a glucose breakdown product inhibits formation of the CAP-cAMP complex required for RNA polymerase to attach at the lac promoter site.

      So yes, suppressor (glucose breakdown product) binds to the cap-cAMP complex so not cannot attach lac promoter site.

      https://www.ncbi.nlm.nih.gov/books/NBK22065/


      It is catabolite repression.
      (1 vote)
  • leafers sapling style avatar for user marie
    are there still sigma factors involved in recruting the RNA polymerase to the promotor?
    (1 vote)
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  • blobby green style avatar for user nurul  ain
    what happens to the metabolism of laactose if there was a mutation in the promoter and operator region?
    (1 vote)
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