EXERCISE 8

REGULATION OF GENES AND THEIR PRODUCTS

Genes determine the nucleotide sequence of RNA molecules. If the RNA is mRNA,
it in turn determines the sequence of amino acids in protein molecules. Thus a gene that
is transcribed into RNA is said to be turned on and the message is translated into specific
protein molecules. The overall process by which genetic information flows from genes to
protein (from the genotype to the phenotype) is called gene expression. The turning on
and off of transcription is the main way that gene expression is regulated. The control of
gene expression makes it possible for cells to produce specific kinds of proteins when
and where they are needed.

Regulation of Gene Action in Prokaryotes

There are two major modes of regulating gene expression in prokaryotes. First is
by controlling the activity of pre-existing enzymes, which is typically a rapid process that
occurs in a matter of seconds. The second mode of regulation, which is a relatively slow
process (few minutes), can be achieved either at the level of transcription or translation.
There are two main types of transcriptional control systems:

A. Negative Transcriptional Control System

- regulatory gene codes for a repressor protein which inhibits transcription of the
structural genes

B. Positive Transcriptional Control System

- regulatory gene codes for an expressor protein which is necessary for the
structural genes to be expressed

Negative control of transcription is possible through enzyme induction or repression.

A. Lactose Operon: Negative Inducible Transcriptional Control System

An operon is a cluster of genes with related functions that exist only in prokaryotes.
The main reason for the clustering of these related genes is for easy coordination of the
said genes. When E. coli is grown on medium containing lactose, all the enzymes needed
for its use are made at once because the operon’s genes are all controlled by a single
switch, the operator. The operator determines whether RNA polymerase can attach to the
promoter and start transcription of the structural genes.

When lactose is absent in the medium, the operon is turned “off”’ by a molecule
called a repressor, a protein that functions by binding to the operator, thereby blocking
the attachment of RNA polymerase to the promoter. The regulator, a gene outside the
operon codes for the repressor. The regulator gene is expressed continually so the cell
always has a supply of repressor molecules and the operon is turned “off”.

The operon is turned “on” when lactose is provided to the medium. Lactose
interferes with the attachment of the repressor to the promoter by binding to the repressor,
changing its shape and making it an in active repressor. With the repressor inactivated,
the repressor cannot bind to the operator. The operator is thus turned “on” since the RNA
polymerase can now bind to the promoter to initiate transcription of the structural genes
that code for the three enzymes needed for lactose utilization. When lactose is totally
consumed, the operon is again turned “off” since there will be no more lactose to
inactivate the repressor, hence it can again bind to the operator preventing the
transcription of the structural genes.

B. The Tryptophan Operon: Negative Repressible Transcriptional Control

System

The tryptophan operon is another type of operon present in prokaryotes. Unlike

the lactose operon, the tryptophan operon is controlled by a repressor that is inactive
when it is alone and is activated only when it is combined with tryptophan. Ordinarily the
operon is turned “on” and the bacterial cell can make tryptophan using enzymes encoded
by the structural genes in the tryptophan operon. However, when tryptophan is provided
in the medium, the cell will stop making tryptophan and simply absorb it from the
surrounding medium. The tryptophan that is provided to the growing medium combines
with the inactive repressor, thereby activating it and allowing the now active repressor to
bind to the operator to prevent transcription of the structural genes.

Procedure:

Each group will construct a model of the lactose and tryptophan operon using
whatever materials are available. The group assigned to conduct the post-lab discussion
will demonstrate/present in class how the model works to illustrate the “turning on or off”
of the operon. Each model should be complete with regards to the structural and
functional components that should be present in the operon.
 WORKSHEET FOR EXERCISE 8
REGULATION OF GENES EXPRESSION IN PROKAYOTES

I. Cite the objectives of the exercise.

II. Demonstration in class of how the structural genes of the lactose and
tryptophan operon are regulated using the constructed models.

III. Guide questions:

1. For each of the following genotypes, explain how mutation (identified by a (–) will
affect the organism grown in lactose medium. Indicate whether
a) B-galactosidase will be synthesized or not,
b) synthesis of B-galactosidase is inducible (I) or constitutive (C) and
c) growth of the organism will occur or not.

a. I-P+O+Z+Y+A+
b. I+P-O+Z+Y+A+
c. I+P+O-Z+Y+A+
d. I+P+O-Z+Y+A+/I+P+O+Z-Y-A-
e. I-P+O+Z+Y+A+/I+P-O+Z+Y+A+

2. Briefly describe how the lactose operon is positively regulated.

3. Indicate what proteins are bound to the regulatory region of the trp operon given
the following conditions. Indicate also if the operon will be turned “on” or “off”.

a. Tryptophan and lactose are present in the medium.

b. Tryptophan and glucose are both absent in the medium.
c. Tryptophan is present but glucose is absent in the medium.
d. Tryptophan and glucose are both present in the medium.