Transcript Document

LECTURE
CONNECTIONS
16 | Control of Gene Expression
in Prokaryotes
© 2009 W. H. Freeman and Company
Genes and Regulatory Elements
• Structural genes: encode proteins that are used in
metabolism or biosynthesis or that play an structural role in
the cell.
• Regulatory genes: encoding products that interact with
other sequences and affect the transcription and translation of
these sequences
• Regulatory elements: DNA sequences that are not
transcribed but play a role in regulating other nucleotide
sequences
Genes and Regulatory Elements
• Constitutive expression: continuously expressed
under normal cellular conditions (encode essential
cellular functions)
• Positive control: stimulate gene expression
• Negative control: inhibit gene expression
Level of Gene Regulation
Genes can be regulated at a number of points along the
pathway:
- Alteration of gene structure (DNA methylation/ changes in
chromatin)
- Transcription
- mRNA processing
- RNA stability (rate mRNA is degraded)
- Translation
- Posttranslational modification
DNA-Binding Proteins
• Much of gene regulation is accomplished by
proteins that bind to DNA sequences and affect
their expression.
• These regulatory proteins generally have discrete
functional parts, called domains.
• Domains: 60 ~ 90 amino acids, responsible for
binding to DNA, forming hydrogen bonds with
DNA
DNA-Binding Proteins
• Distinctive types of DNA-binding proteins based on the
motif
• Motif: within the binding domain, a simple structure
that fits into the major groove of the DNA
Concept Check 1
How do amino acids in DNA-binding proteins
interact with DNA?
a. By forming covalent bonds with DNA base
b. By forming hydrogen bonds with DNA base
c. By forming covalent bonds with sugars
Concept Check 2
What is the difference between a structural gene and a
regulator gene?
a. Structural genes are transcribed into mRNA, but regulator
genes are not.
b. Structural genes have complex structures; regulator genes
have simple structure.
c. Structural genes encode proteins that function in the
structure of the cell; regulator genes carry out metabolic
reactions.
d. Structural genes encode proteins; regulator genes control
the transcription of structural genes.
16.3 Operons Control Transcription in
Bacterial Cells
• A group of bacterial structural genes that are transcribed
together (along with the promoter and additional
sequences that control transcription) is called an
operon.
• Operon: promoter + additional sequences that control
transcription (operator) + structure genes
• Regulator gene: DNA sequence encoding products that
affect the operon function, but are not part of the operon
Negative and Positive Control; Inducible and
Repressible Operons
• Inducible operons: Transcription is usually off and
needs to be turned on.
• Repressible operons: Transcription is normally on
and needs to be turned off.
Negative and Positive Control; Inducible and
Repressible Operons
• Negative inducible operons: The control at the
operator site is negative.
The binding of the regulator protein to the operator inhibit
transcription.
Such operons are usually off and need to be turned on by an inducer- so the transcription is called inducible.
• Inducer: small molecule that binds to the repressor and
turns on the transcription
Negative and Positive Control; Inducible and
Repressible Operons
• Negative repressible operons: Some operons with
negative control are repressible, meaning that
transcription normally takes place and must be turned
off.
• Corepressor: a small molecule that binds to the
repressor and makes it capable of binding to the
operator to turn off transcription.
Negative and Positive Control; Inducible and
Repressible Operons
• With positive control, a regulatory protein is an activator:
it binds to DNA and stimulates transcription.
• Positive inducible operon: transcription is normally
turned off because the regulator protein (an activator) is
produced in an active form.
• Positive repressible operon: transcription normally
takes place and has to be repressed. Transcription is
inhibited when a substance besomes attached to the
activator and renders it unable to bind to DNA.
Concept Check 3
In a negative repressible operon, the regulator protein is
synthesized as
.
a.
b.
c.
d.
an active activator
an inactive activator
an active repressor
an inactive repressor
The lac Operon of E. coli
• A negative inducible operon
• Inducer: allolactose
• lacI: repressor encoding gene
• lacP: operon promoter
• lacO: operon operator
The lac Operon of E. coli
• Structural genes
• lacZ: encoding β-galactosidases
• lacY: encoding permease
• lacA: encoding transacetylase
• The repression of the lac operon never completely shuts
down transcription.
The repression never completely shuts down.
Concept Check 4
In the presence of allolactose, the lac repressor
_________ .
a.
b.
c.
d.
binds to the operator
binds to the promoter
cannot bind to the operator
binds to the regulator gene
lac Mutations
•
Partial diploid: full bacterial chromosome + an extra
piece of DNA on F plasmid
•
Structural gene mutations: affect the structure of the
enzymes, but not the regulations of their synthesis
Partial diploids with lacZ+ and lacY− on the bacterial
chromosome and lacZ− and lacY+ in the plasmid
functioned normally producing β-galactosidase and
permease.
lac Mutations
• Regulator gene mutations: lacI− leads to constitutive
transcription of three structure genes.
• lacI+ is dominant over lacI− and is trans acting. A single
copy of lacI+ brings about normal regulation of lac
operon.
• lacI+lacZ−/lacI−lacZ+ produce fully functional βgalactosidase.
lac Mutations
• Operator mutations: lacOc (C = constitutive)
• lacOc is dominant over lacO+, which is cis acting.
Cis-acting: able to control the expression of genes only
when on the same piece of DNA only.
Trans-acting: able to control the espression of genes on
other DNA molecules.
• lacI+ lacO+ lacZ- and lacI+ lacOc lacZ+ produce fully
functional β-galactosidase constitutively.
(If the operator is mutated, the repressors can not bind to it!)
lac Mutations
• Promoter mutations
• lacP−: cis acting
• These mutations interfere with the binding of RNA
polymerase to the promoter.
• lacP- mutations don’t produce lac enzymes either in
the presence or in the absence of lactose.
Positive control and catabolite repression
• Catabolite repression: using glucose when available,
and repressing the metabolite of other sugars.
• This is a positive control mechanism: The positive
effect is activated by catabolite activator protein (CAP).
cAMP is binded to CAP, together CAP–cAMP complex
binds to a site slightly upstream from the lac gene
promoter.
Positive control and catabolite repression
• cAMP – adenosine-3′,5′-cyclic monophosphate
• The concentration of cAMP is inversely proportional to
the level of available glucose.
Concept Check 4
What is the effect of high levels of glucose on the lac
operon?
a.
b.
c.
d.
Transcription is stimulated.
Little transcription takes place.
Transcription is not affected.
Transcription may be stimulated or inhibited, depending on
the levels of lactose.
The trp Operon of E. coli
• A negative repressible operon
• Five structural genes
• trpE, trpD, trpC, trpB, and trpA – five enzymes
together convert chorismate to tryptophane.