The document discusses the galactose and histidine operons. It describes the structural organization and regulation of the galactose operon, including the Leloir pathway for D-galactose metabolism and repression/induction by galactose. It also summarizes the structural organization and two mechanisms of regulation (feedback inhibition and repression control) for the histidine operon. Finally, it explains attenuation control of the histidine operon through transcriptional termination or anti-termination depending on histidine availability.

1. BANGALORE UNIVERSITY
JB CAMPUS BANGALORE
DEPARTMENT OF MICROBIOLOGY, BIOTECHNOLOGY & FOOD
TECHNOLOGY
TOPIC : GALACTOSE & HISTIDINE OPERON
GUIDED BY : Dr. RAVIKIRAN.T PRESENTED BY : PUNITH KUMAR.S
DEPT.OF BIOTECHNOLOGY M.Sc II Sem. BIOTECHNOLOGY
BANGALORE UNIVERSITY

2. Contents :
Operon
Galactose operon
Structural organization
Metabolism
Regulation
Histidine operon
Structural organization
Regulation
Conclusion
Reference

3. OPERON
An operon is a group of closely linked genes (structural and regulatory
gene),which regulate the metabolic pathway in prokaryotes ,but not in
eukaryotes.
In eukaryotes, each gene is made up of an individual mRNA and each gene
has its own promoter .
A bacterium contains thousands of gene, when all genes functions at a
same time the use will be flooded with enzymes and proteins. So, the genes
for the required enzyme at that particular time are switched on and the
other genes are switched off.
This on and off mechanism is explained by the operon model.
HISTORY : Operon were first identified as mode of expression control in
1961by Francois Jacob and Jacques Monod .

4. Galactose operon
 Gal operon is a prokaryotic operon.
Codes enzymes for an amphibolic pathway of D-galactose metabolism.
Gal operon performs catabolic and Anabolic function
Catabolic Function : Galactose is used as Carbon source
Catabolic Function : UDP Galactose UDP glucose
 Anabolic Function : UDP glucose UDP galactose
This leads to the formation of lipopolysaccharide (cell wall component)

5. Structural organization of operon

6. Structure
 Operator 2
OE = Operator external
OI = Operator internal
 Promoter 2 Overlapping
P1 & P2 = Transcription start site
 hbs = histone unit binding site
Activator sequence(AS)
Structural genes 4
Gal E = UDP – galactose – 4 – epimerase
Gal T = galactose – 1 – phosphate uridylyl transferase
Gal K = galactokinase
Gal M = mutarotase

7. This operon when performs catabolism.
Galactose imported into the cell by permease
Or galactose formed intercellularly by disaccharide metabolism
In galactose metabolism , β – D galactose is not utilized, thus its
converted into α-D galactose.

8. Leloir pathway of D-Galactose metabolism

9. REGULATION OF GAL OPERON
1) Gal R mediated DNA loop formation / repression
Gal R gene encode tetrameric repressor that binds to 2 operators one located
internally and one located externally
This process requires presence of histone like protein (HU) and supercoiled
DNA with operators in same face.
Interaction of two operator bound Gal R results in formation of DNA loop
(represosome)
Looping of DNA blocks the access of RNA polymerase to promoters or
inhibits formation of open complex.
Structural genes are not transcribed.

10. 2) Gal R binds a dimer to OE
A) Inhibits Promotor P1 ( transcription off )
Inhibits formation of open promotor complex through RNA polymerase contact
with α CTD
Also causes CONTACT INHIBITION OF P1.

11. B) Activates promoter P2 ( transcription on )
Allows low levels structural genes to be expressed
NOTE: The Gal R – OI complex will not cause any repression because of
presence of ANTI-PAUSE sequence in intermediate upstream area.

12. 3) Induction of Gal Operon by D - galactose
Condition – 1
Increased level of D – galactose ( both α & β ) in medium
Bind to allosteric site of repressor ( dimer )
Resulting in conformational change of protein repressor
Suppress interaction of repressor with RNA P and DNA
Induce operon and increase rate of galactose metabolism.

13. Condition – 2 : DNA looping
D-galactose first breaks Gal R – Gal R tetramers into ,
Individual Gal R – OE and Gal R – OI dimer ( P1 is repressed and P2 is
activated )
Gal R – OI ( dissociated by D – Galactose )
Gal R – OE ( dissociated and P1 is activated )

14. 4) Regulation of CRP-CAMP complex ( CCC)
CRP-cAMP complex binds at activation site (AS)
i. Blocks RNA polymerase to form open complex with P2 ( Transcription stops )
ii. CRP-cAMP leads to formation of closed complex at P1 with RNA polymerase that further
isomerize to open complex ( Transcription ON )
NOTE :
When glucose concentration is high in medium CRP-cAMP concentration will be low
Basal level transcription occurs at P2 by formation of binary complex
cAMP and its receptor protein CRP complex enhances P1 but represses P2

15. HISTIDINE OPERON
Histidine operon and its regulation of action in the bacterium Salmonella
typhimurium.
The bacteria controls its rate of histidine biosynthesis by 2 ways:
1. Intracellular concentration rises and feedback inhibition shuts down the
pathway
When histidine is being transported into the cell, the intracellular concentration
rises and feedback inhibition shuts down the pathway.
When external histidine is exhausted , the histidine pool fails until feedback
inhibition is retrieved and biosynthesis of the amino acid is resumed .

16. 2. Repression control, Governs the intracellular concentration of
biosynthetic enzymes.
This method is considerably slower in action than feedback inhibition although
the enzymes concentration may begin new steady state level is reached.
Presence or absence of external histidine is not the only factor regulating the
rate of histidine biosynthesis.
The bacterium also keeps its rate of synthesis of amino acid in line with its
growth rate.

17. Structural organization of operon
Histidine operon contains cluster of 9 structural gene his G,D,C,B,H,A,F,I,E which codes for
enzymes aids in the pathway of synthesis of histidine from 5-phosphoribosyl 1-
pyrophosphate(PRPP).
There are 5 regulatory genes viz. his R,U,S,T and W that are associated with the operon but not
closely linked to it .
The gene his R codes for histidine tRNA , while his S codes for histidyl-tRNA synthetase.
The histidine operon lacks an operator region and a CRP site.

18. Because of this lack of an operator region , the transcriptional control in histidine operon solely
takes by attenuation.
The transcription produces a single polycistronic mRNA that is about 7300 nucleotides long ,
extending from primary promoter to Rho-independent terminator.
2 week internal promoters , Hisp2 and Hisp3 are located within the HisC and HisF genes
respectively.

19. Regulation : Control of Transcription Initiation
& Elongation
Transcription of the his operon is about four fold more efficient in bacteria
growing in minimal glucose medium than when growing in rich medium.
This form of control is called metabolic regulation , adjusts the expression of the
operon to the amino acid supply in the cell.
It is mediated by “alarmone” (ppGdd) guanosine 5’-diphosphate 3’-diphosphate ,
which is the effector of the stringent response.
The alarmone regulates the his operon positively by stimulating the primary
promoter hisp1 under conditions of moderate amino acid starvation.

20. Regulation : Attenuation control of the histidine
operon
Attenuation is the modification of gene expression by events that influence
aspects of transcription other than initiation of transcription .
The attenuator is located in the leader region of the RNA transcript that lies
between the promoter and the 1st structural gene.
It comprises a segment coding for the leader peptide followed by a terminator
sequence .

21. The his-specific regulatory element is transcribed in a 180-nucleotide RNA leader
, which exhibits two prominent features:
(i) A 16-residue coding sequence in the leader peptide –coding region ,
including seven consecutive codons specifying histidine .
The amino acid sequence is :
(ii) overlapping regions of dyad symmetry capable of folding into mutually
exclusive, alternative secondary structure that signal either transcription termination
or anti termination. There are 3 stems in the termination configuration and 2 in the
antitermination configuration.

22. Negative regulation : RNA Termination
Translation control of His operon, transcription is determined by ribosomes
occupancy of leader RNA which in turn depends given the peculiar composition of His leader
peptide on the availability of His tRNA .
When histidine is present in sufficient amount, the supply of histidyl tRNA is plentiful.
This permits the translating ribosome to pass across the leader sequence of RNA without pausing.
The relevant codons and adjacent nucleotides are free to base pair , resulting in the formation of
the RNA terminator this prevents the translation of his operon structural genes.

23. Negative regulation : RNA Termination
In case of severe limitation of intracellular pool of all charged tRNAs,
translation of leader peptide fails to initiate: under these condition, the A:B, C:D,
E:F stem loop structures from sequentially, producing a strong transcription
termination.

24. Positive regulation : Anti-termination
When histidine is in low concentration , the supply of histidyl tRNA is limited,
this result in the ribosome pausing or stalling as it passes across the leader
sequence and masking the codons, this permits the base downstream to base pair
and from the antitermination structure , which in turn prevents the formation of
the terminator.
Translation of the his operon structural genes therefore takes place.

25. Conclusion
The operon , genetic regulatory system found in prokaryotes ,in which genes
coding for functionally related proteins are clustered along the DNA. This
feature allows controlled protein synthesis according to the needs of cell.
The gal operon of E.coli plays a major role in cellular metabolism by encoding
enzymes that catalyze conversion of D-galactose to energy sources as well as
to anabolic substrates.
The histidine operon have served as powerful model system for studying
fundamental evolutionary , metabolic ,physiological and genetic processes ,
such as gene duplication , transposition , mutagenesis, including the widely
used Ames test.

26. Reference
Books
John S. Kovach. Robert F. Goldberger. Journal of bacteriology. 2006. volume 97. page no : 1283-
1290.
Chang GW, Roth jr, Ames BN (2009). Histidine regulation in salmonella typhimurium.
8.mutation of his T gene. J Bacteriol 108:410-414
Journal
Irani M.H, Orosz L, Adhya S(1983). A control element within a structural gene; The gal operon
of Escherichia coli.Cell 32(3):783-788.https://doi.org/10.1016/0092-8674(83)90064-8.
Dale E A Lewis , Sankar Adhya(2015). Molecular mechanism of transcription initiation at gal
promoters and their multi-level regulation by GalR, CRP and DNA loop.Biomolecules.5(4):2782-
807.https://doi.org/10.3390/biom5042782.

27. THANK YOU