Operons

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Operons

• Charmaine Robinson

GGG 291 Fall 2008
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Nobel Prize 1965
for their discoveries concerning genetic
control of enzyme and virus synthesis
François Jacob
André Lwoff
Jacques Monod
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François Jacob (b. 1920)

• Attended Lycée Carnot in Paris
• Studied medicine at the Faculty of Paris
• - surgeon
• 1940 Free French Forces (London)
• Wounded during WWII, serving in Africa
• Second Armoured Division
• - severely wounded in Normandy 1944
• Awarded the Croix de la Libération
• Completed medical degree in 1947
• Science degree in 1951
• Doctorate in 1954 at Sorbonne (lysogenic
  bacteria, provirus)
• 1950 joined the Institut Pasteur under Dr. Lwoff
• Mostly studied bacteria, bacteriophages
• 1958 w/Monod discovered the operon

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Jacques Monod (1910- 1976)

• 1931 Faculty of Paris, degree in Natural
  Sciences
• 1934 Lecturer at Faculty of Sciences
• 1936 Rockefeller grant at CA Institute of
  Technology (in Morgans lab)
• 1941 doctorate in Natural Sciences
• Joined Institut Pasteur in Lwoffs dept.
• 1958 discovered the operon w/Jacob
• 1971 appointed Director of Institut Pasteur

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How the Operon was discovered Genetic
regulatory mechanisms in the synthesis of
proteins
IF A given agent specifically alters the rate of
synthesis of a protein THEN 1.) Does the agent
act at the cytoplasmic level or at the genetic
level? 2.) Does the specificity depend on some
feature of the transfer of information from gene
to protein, or upon another element (not
represented in the structure of the protein,
gene, or messenger)?
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Used Enzymatic systems and Lysogenic systems
WHY? 1.) Defined systems where the synthesis of
a protein is seen to be controlled by specific
agents 2.) ß-galactosides initiating
biosynthesis at one end of the corridor, and UV
light on lysogenic bacteria initiating
biosynthesis at the other end
Pasteur Institute in France
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Kinetics of Induction using galactosidase

• Found a linear relationship between enzyme
  activity and total protein
• Enzyme formation is FAST after addition of
  inducer (3 min!)
• Enzyme synthesis stops as quickly with removal
  of inducer
• CONCLUSION
• Inducer does NOT activate synthesis of a stable
  intermediate

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Specificity of Induction using galactosidase

• Tested inducers of galactosidase and substrates
• No correlation between affinity for the enzyme
  and capacity to induce
• CONCLUSION
• Mechanism of induction does not imply any
  inherent correlation between molecular structure
  of the inducer and structure of the binding site
  (on enzyme).

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Enzyme Repression

• Important to distinguish inhibition of enzyme
  synthesis, and not inhibition of activity.
• Tested Kinetics and Specificity of repression
• CONCLUSIONS
• Both repression and induction appear to control
  rate of synthesis of proteins in a similar
  manner.
• Different enzymes, with related functions, are
  frequently co-induced or co-repressed by a single
  agent.

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Regulator Genes (Mutants of LacZ)

• Three distinct genes for the Lac system
• - ß-galactosidase (Z)
• - permease (Y)
• - regulator (makes the repressor) (I)
• Isolated mutants of each gene
• Both haploids, and diploids by crosses using
  the F-factor
• Studied the phenotypes of these mutants

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Regulator Genes (Mutants of LacZ)
RESULTS

• 1.) Constitutive i mutants showed repressor acts
  in trans
• (i mutant allele can be located on F chromosome
  or the bacterial chromosome, in a diploid
  heterozygote)
• 2.) Constitutive mutant alleles of z and y must
  be cis with the operator
• 3.) Constitutive operator mutants are dominant
  (even the heterozygote drives expression)

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Regulator Genes and phage systems
Phage life cycles Lysis (vegetative stage)
replication, production of infectious
phage Lysogeny (prophage stage) phage genome
attaches to bacterial genome, stays quiescent, no
new phage
Lambda phage
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Regulator Genes and phage systems

• Used UV, X-rays, and other compounds to induce
  lysogenic bacteria to produce phage (lysis)
• Examined these mutants to determine how the
  prophage inhibits phage replication
• - either chromosomal
• - or cytoplasmic
• CONCLUSIONS
• Expression of viral (phage) genes is controlled
  by a cytoplasmic repressor, which is synthesized
  by one regulator gene.

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Messenger RNA, candidate for the intermediate
in protein synthesis
Characteristics of the intermediate 1.) a
polynucleotide 2.) heterogeneous in molecular
weight 3.) base composition reflects composition
of DNA 4.) associated with ribosomes
mRNA!
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CONCLUSIONS
Basic Components of Operons (as defined by Monod
and Jacob) 1.) governs expression of several
closely linked genes 2.) is adjacent to those
genes 3.) sensitive to a repressor
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Big Picture

• Cells not only have genetic plans for proteins,
  ALSO have genetic regulatory system for
  expressing those plans!
• Discovery changed how people viewed development
  in higher organisms.

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Discussion Points

• How much of these discoveries are due to being
  at the right place at the right time, plus
  working on the right model organism?
• How many of you became interested in Genetics
  (or science in general) from reading one paper or
  attending one course/lecture?
• Random fact Monod talks about using a strain
  of E. Coli that had been isolated from Lwoffs
  intestine tract.(!)