CHP 16 - PROKARYOTES
Recap
- DNA is arranged into genes
- these genes provide storage of info
- this info is expressed through transcription & translation
Efficient expression of genetic info is dependent on control mechanisms
promote ↲ ↳ suppress
Gene expression in prokaryotes
Early studies in bacteria & yeast: easy to culture & mutate, noticed fluctuating protein levels in response to
environmental conditions
Adaptation hypothesis: lactose in growth medium induces expression of enzymes specific for lactose
metabolism
Constitutive, inducible, repressible systems, negative or positive control:
- constitutive genes – permanently expressed, regardless of environment ∴ “housekeeper”
- inducible – expressed in response to a condition (e.g. inducer – lactose)
- negative control – expression (transcription) happens unless actively suppressed (must be switched off)
- positive control – expression is suppressed unless actively stimulated (must be switched on)
Inducible expression of lactose operon E. Coli
- lactose (lac) = galactose + glucose
- inducible & lactose = the inducer
↳ the inducer is allolactose - isomer of lactose
Lac operon
Lac operon: A set of genes that are specific for uptake & metabolism of lactose E. coli & other bacteria.
↳ consists of 3 structural genes & 2 regulatory regions upstream (cis & trans action)
Operon: An operon is a cluster of genes that are transcribed together to give a single messenger RNA
(mRNA) molecule, which ∴ encodes multiple proteins
Prokaryotic genes with related functions are organized in groups & are expressed in a coordinated fashion
(operon)
Structural genes:
lacZ – b-galactosidase (respons. for converting lactose to glucose & galactose)
lacY – b-galactoside permease (facilitates the entry of lactose into the cell)
lacA – transacetylase (role not clear, maybe involved in removal of toxic by-products of lactose digestion
from the cell)
- polycistronic mRNA means that the structural genes code for several different proteins
, How does lactose regulate transcription of the three structural genes?
- Gene activity is repressed when Lactose (lac) is absent, because Lac is the inducer of gene expression
- Lactose present – we see an induced amount of gene expression
- No lactose present/absent – there is less expressed or none expression
- induction is not mediated through interaction with enzymes: Isopropyl β-d-1- thiogalactopyranoside
(IPTG) is a molecular biology reagent
- this compound is a molecular mimic of allolactose, a lactose metabolite that triggers transcription of the
lac operon, & it is ∴ used to induce protein expression where the gene is under the control of the lac
operator
- constitutive mutations – lacI- & lacOc in repressor (allosteric) & operator regions. lacI encodes an
allosteric repressor molecule
lac repressor protein: an allosteric protein which has 2 forms:
- an active form in which the repressor bind to the lac operator & blocks transcription of the lac operon
structural genes when lactose is absent
- an inactive form which cannot bind the lac operator. The inactive form is dominant when lactose is
present & the inducer allolactose binds to the lac repressor protein
- repressor binds to operator region – inhibits RNA polymerase binding
- repressor gene (I) encodes for lactose-
binding site (white)
- RNA pol (3 teeth) binds to the promoter (P)
- lactose is present, ∴ repressor gene is not
applicable, ∴ transcription takes place
I – repressor
Superscripts: P – promoter
- plus sign (+) = gene is in wildtype/normal form O – operator
- neg sign (-) = gene has been mutated L - leader
Recap
- DNA is arranged into genes
- these genes provide storage of info
- this info is expressed through transcription & translation
Efficient expression of genetic info is dependent on control mechanisms
promote ↲ ↳ suppress
Gene expression in prokaryotes
Early studies in bacteria & yeast: easy to culture & mutate, noticed fluctuating protein levels in response to
environmental conditions
Adaptation hypothesis: lactose in growth medium induces expression of enzymes specific for lactose
metabolism
Constitutive, inducible, repressible systems, negative or positive control:
- constitutive genes – permanently expressed, regardless of environment ∴ “housekeeper”
- inducible – expressed in response to a condition (e.g. inducer – lactose)
- negative control – expression (transcription) happens unless actively suppressed (must be switched off)
- positive control – expression is suppressed unless actively stimulated (must be switched on)
Inducible expression of lactose operon E. Coli
- lactose (lac) = galactose + glucose
- inducible & lactose = the inducer
↳ the inducer is allolactose - isomer of lactose
Lac operon
Lac operon: A set of genes that are specific for uptake & metabolism of lactose E. coli & other bacteria.
↳ consists of 3 structural genes & 2 regulatory regions upstream (cis & trans action)
Operon: An operon is a cluster of genes that are transcribed together to give a single messenger RNA
(mRNA) molecule, which ∴ encodes multiple proteins
Prokaryotic genes with related functions are organized in groups & are expressed in a coordinated fashion
(operon)
Structural genes:
lacZ – b-galactosidase (respons. for converting lactose to glucose & galactose)
lacY – b-galactoside permease (facilitates the entry of lactose into the cell)
lacA – transacetylase (role not clear, maybe involved in removal of toxic by-products of lactose digestion
from the cell)
- polycistronic mRNA means that the structural genes code for several different proteins
, How does lactose regulate transcription of the three structural genes?
- Gene activity is repressed when Lactose (lac) is absent, because Lac is the inducer of gene expression
- Lactose present – we see an induced amount of gene expression
- No lactose present/absent – there is less expressed or none expression
- induction is not mediated through interaction with enzymes: Isopropyl β-d-1- thiogalactopyranoside
(IPTG) is a molecular biology reagent
- this compound is a molecular mimic of allolactose, a lactose metabolite that triggers transcription of the
lac operon, & it is ∴ used to induce protein expression where the gene is under the control of the lac
operator
- constitutive mutations – lacI- & lacOc in repressor (allosteric) & operator regions. lacI encodes an
allosteric repressor molecule
lac repressor protein: an allosteric protein which has 2 forms:
- an active form in which the repressor bind to the lac operator & blocks transcription of the lac operon
structural genes when lactose is absent
- an inactive form which cannot bind the lac operator. The inactive form is dominant when lactose is
present & the inducer allolactose binds to the lac repressor protein
- repressor binds to operator region – inhibits RNA polymerase binding
- repressor gene (I) encodes for lactose-
binding site (white)
- RNA pol (3 teeth) binds to the promoter (P)
- lactose is present, ∴ repressor gene is not
applicable, ∴ transcription takes place
I – repressor
Superscripts: P – promoter
- plus sign (+) = gene is in wildtype/normal form O – operator
- neg sign (-) = gene has been mutated L - leader
