GENE REGULATION - TRP OPERON

1. GENE REGULATION IN
PROKARYOTES
S.PALANIANANTH 1ST MSC MICROBIOLOGY
AYYA NADAR JANAKI AMMAL COLLEGE SIVAKASI

2. TRP OPERON
SUBMITTED TO : DR. S. MUNIASAMY
ASSISTANT PROFESSOR OF MICROBIOLOGY
AYYA NADAR JANAKI AMMAL COLLEGE SIVAKASI

3. • Gene regulation
 Gene regulation : It is the process by which information from a gene is used in the synthesis of a
functional gene product . These products are often proteins, but in non-protein coding genes such as
rRNA genes or tRNA genes, the product is a functional RNA.
 Types of regulation of gene expression
 Positive regulation or induction :
 When the expression of genetic is quantitatively increased by the presence of specific regulatory element is
known as positive regulation.
 Element modulating positive regulation is known as inducer, activator or positive regulator .
 Negative regulation or repression.
 When the expression of genetic information diminished by the presence of specific regulatory element.
 The element or molecule mediating the negative regulation is said to be repressor Activate

4. Generalized structure of operon
 Promoter:
 A promoter is a nucleotide sequence in the DNA that initiates transcription of a particular gene.
Operators :
These are segments of DNA that regulate the activity of the structural genes of the operon
.• Structural Genes:
A gene that codes for any RNA or protein product other than a regulatory factor
• Regulator
These nucleotide sequences control the operator gene in cooperation with certain compounds called
inducers and co-repressors present in the cytoplasm.

5. Operon
 The operon, principle of gene expression
 An operon is a functioning unit of genomic DNA containing a cluster of genes under the control of a single
promoter.
 In bacteria, genes that encode for proteins with closely related functions are found grouped along with cis-
acting regulatory elements that determine the transcription of these genes, thus these genes are regulated in
a coordinated way. These clusters of genes are called operons. The transcription product of an operon is a
single polycistronic MiRNA
Types of operon
Inducible operons:
They include genes that encode for enzymes that take part in metabolic pathways.The expression of the
gene is controlled by the substrate. Example is the "Lac Operon“
Repressible operons:
They include genes that encode for enzymes involved in biosynthetic pathways.The expression of the
gene is controlled by the end-product of the pathway. Example is the "Trp Operon".

6. INTRODUCTION
 The trp operon is an operon a group of genes that is used , or transcribed that codes for the
components for production of tryptophan.
 The trp operon is present in many bacteria but was first characterized in E.coli.
 Trp operon discovered in 1953 by Jacques Monod and colleagues , the trp operon in E.coli was the first
repressible operon to be discovered.
 Tryptophan is an aromatic amino acid, and is required for synthesis of all protein that contain
tryptophan. If tryptophan is not present in the medium in adequate quantity, the bacterial cell has to
make it, as it is required for the growth of the bacteria.
 The tryptophan operon, found in E.coli bacteria, it is a group of genes that encode biosynthetic
enzymes for the amino acid tryptophan (Repressible OPERON)
 The tryptophan operon is expressed (turned on) when tryptophan levels are low and repressed (turned
off) when they are high

7. INTRODUCTION
 The trp operon is regulated by the trp repressor. When bound to tryptophan, the trp repressor blocks
expression of the operon.
 Tryptophan biosynthesis is also regulated by attenuation (a mechanism based on coupling of
transcription and translation).

8. What is the trp operon?
 Bacteria such as Escherichia coli (a friendly inhabitant of our gut) need amino acids to survive—
because, like us, they need to build proteins. One of the amino acids they need is tryptophan.
 If tryptophan is available in the environment, E. coli will take it up and use it to build proteins.
However, E. coli can also make their own tryptophan using enzymes that are encoded by five genes.
These five genes are located next to each other in what is called the trp operon.
 If tryptophan is present in the environment, then E. coli bacteria don't need to synthesize it, so
transcription of the genes in the trp operon is switched "off." When tryptophan availability is low, on
the other hand, the operon is switched "on," the genes are transcribed, biosynthetic enzymes are
made, and more tryptophan is produced.

9. Structure of the trp operon

10. Structure of the trp operon
 Tryptophan operon consists of structural gene and regulatory gene.
 I. Regulatory gene are Promoter, Repressor, Operater and Leader sequence.
 II. Structural gene are TrpE, TrpD, TrpC, TrpB and TrpA
 trpE: It enodes the enzyme Anthranilate synthase I
 trpD: It encodes the enzyme Anthranilate synthase II
 trpC: It encodes the enzyme N-5’-Phosphoribosyl anthranilate isomerase and Indole-3-
glycerolphosphate synthase
 trpB: It encodes the enzyme tryptophan synthase-B sub unit
 trpA: It encode the enzyme tryptophan synthase-A sub unit


11. Turning the operon "on" and "off"
 What does the operator do?
 This stretch of DNA is recognized by a regulatory protein known as the trp repressor. When the repressor
binds to the DNA of the operator, it keeps the operon from being transcribed by physically getting in the way
of RNA polymerase, the transcription enzyme.
 The trp repressor does not always bind to DNA. Instead, it binds and blocks transcription only when tryptophan is
present. When tryptophan is around, it attaches to the repressor molecules and changes their shape so they
become active. A small molecule like tryptophan , which switches a repressor into its active state, is called
a corepressor.

12. Tryptophan operon is regulated by following mechanism
1. Repression
 When there is little tryptophan in the cell, on the other hand, the trp repressor is inactive (because no
tryptophan is available to bind to and activate it). It does not attach to the DNA or block transcription,
and this allows the trp operon to be transcribed by RNA polymerase.
 In this system, the trp repressor acts as both a sensor and a switch. It senses whether tryptophan is
already present at high levels, and if so, it switches the operon to the "off" position, preventing
unnecessary biosynthetic enzymes from being made.

13. Repression
 When tryptophan is high in cell then it binds with repressor protein and change its confirmation so that
it become active and bind to the operator near promoter.
 Binding of repressor protein to operator overlaps the promoter, so RNA polymerase cannot bind to the
promoter. Hence transcription is halted.
 Since tryptophan is already high in cell, no transcription of structural gene is required for biosynthesis
of tryptophan. This is also known as negative regulation.

14. High tryptophan

15. Tryptophan operon is regulated by following mechanism
2. Attenuation:
 In bacteria, transcription and translation occurs simultaneously. The translation starts before
transcription completes. In this attenuation mechanism, rate of translation determines whether
transcription continues or terminates. Therefore the attenuation mechanism is only found in bacteria
but not in eukaryotic cell.
 It Involves the trpL gene, a sequence of 162 nucleotides that produces an mRNA leader sequence; near
the 3’ end of this leader sequence (trpL gene) is a region called the attenuator.
.

16.  Leader sequence (trpL) play important role in attenuation. Leader sequence contains such a nucleotide
sequence that mRNA transcribed from it contains four specific region. Region 1, region 2, region 3 and
Region 4. Region 3 is complementary to both region 2 and region 4.
 If region 3 and region 4 base pair with each other, they form a loop like structure called attenuator and
it function as transcriptional termination. If pairing occur between region 3 and region 2, then no such
attenuator form so that transcription continues.
 Region 1 is the most important region that determines whether to form loop between region 2-region 3
or region 3-region 4. The region 1 consists of sequence of 14 codons, out of which two codons are
tryptophan codon (codon 10 and 11)

18.  When tryptophan is high in cell than tRNA carrying tryptophan encodes codon 10 and 11. Such that
ribosome encloses the region 2 which is near to the tryptophan codon. Hence region 3 base pair with
region 4 to form attenuator as region 2 is not available for pairing. Consequently, transcription is halted.
 When tryptophan is low or absent in cell, then translation stops at the position of tryptophan codon.
Such that loop between region 2 and region 3 forms. Transcription continues.

19. The biosynthesis of tryptophan

20. The biosynthesis of tryptophan

TRP E-Anthranilate synthetase I Converts chorismate to anthranilate using
ammonia.
 Anthranilate is an amino acid that is later
broken down into another amino acid, Trysoine.

TRP D-Anthranilate synthetase IIA type of Glycosyltransferase that participates
in the synthesis of Tryptophan
 Produces an amino acid called Anthranilate which
,by the help of TRP E, is broken down into
another amino acid, Trysoine.
 Converts -(5-phospho-D-ribosyl)-anthranilate and
diphosphate to anthranilate and
5-phospho-alpha-D-ribose 1-diphosphate.
 It is 1 of the 2 components that together encode
Anthranilate synthetase

21.  TRP C- indole-3-glycerol-phosphate synthase Involved in the synthesis of tryptophan and amino
acids.
 It is a type of lyase that cleaves carbon-carbon
bond.
 this ability allows it to converts
1-(2-carboxyphenylamino)-1-deoxy-D-ribulose
5-phosphate to C(1)-(3-indolyl)-glycerol
3-phosphate and carbon dioxide and water
TRP B tryptophan synthase subunit B There are two beta subunits which form a dimer
 Beta dimer catalyzes the formation of
L-tryptophan from L-serine and 1-(indol-3-yl)glycerol 3-phosphate

22.  TRP A tryptophan synthase subunit A Tryptophan synthase is an enzyme found in plants
and bacteria
 There are two separate alpha subunits
 They catalyze the formation of indole and
glyceraldehyde 3-phosphate from indole glycerol
phosphate in tryptophan biosynthesis, which is
needed by the B subunits.

23. Questions
 What is meant by catabolism and anabolism process?
 TRP OPERON IS ANABOLIC OR CATABOLIC
 Role of RNA polymerase?
 Define Apo enzyme?
 TRP OPERON OR OTHERWISE CALLED?
 ARE OPERONS PRESENT IN EUKARYOTES?
 WHEN WILL TRP OPERON IS TURNED ON? AN TURNED OFF?
 WHAT IS POLYCISTRONIC MRNA?
 HOW MANY STRUCTURAL GENES ARE PRESENT IN THE TRP OPERON ?
 TRP OPERON IS AN EXAMPLE OF ______________________