section 26.2
Gene Regulation in Prokaryotes
595
lac mRNA is not made. This is the negative regulation
of the lac operon. Mutants have been isolated that
produce inactive repressor proteins; these cells make
lac mRNA continuously (constitutive synthesis), even
in the absence of lactose. Actually, some lac mRNA is
always made at a level of about one or two
transcription events per generation because repression
is never complete. This basal level synthesis is
responsible for a very small amount of the proteins.
4. A sequence of bases (in the DNA)—the operator
lac o—is adjacent to the lac mRNA promoter and
binds repressor. When the repressor protein is bound
to the operator, attachment of RNA polymerase to the
promoter is prevented by steric interference, and
initiation of transcription of lac mRNA does not
occur. A mutation in the operator, which eliminates
repressor binding, also leads to constitutive synthesis.
5. An inducer (a small effector molecule) is needed to
initiate transcription of lac mRNA. The inducer
of the lac operon, which is allolactose,
[yd-D-galactopyranosyl-(l -*
6
)-/3 -D-gI ucopyranose ]
a structural isomer of lactose formed by basal
synthesis of /3-galactosidase, binds to the repressor
and alters its three-dimensional structure such that it
is unable to bind to the operator. Thus, in the presence
of lactose or other inducers the operator is unoccupied
and the promoter is available for initiation of mRNA
synthesis. It is common to refer to inactivation of the
repressor by an inducer as derepression.
Thus, the overall regulatory pattern is as follows: A
bacterium growing in a medium without lactose does not
make either the lac z or lac y product because the repressor
that is made is bound to the operator and prevents synthe-
sis of lac mRNA. The cell grows by utilizing whatever
other carbon source is available. If lactose is added (and
if glucose is absent; see below), basal level lac y and lac
z proteins bring the lactose into the cell and convert it to
allolactose. As a result, the repressor is inactivated, RNA
polymerase binds to the promoter, and synthesis of lac
mRNA begins. Lac mRNA synthesis continues until the
lactose is exhausted, in which case the inactive repressor
would be reactivated and repression reestablished.
The lac operon is also positively regulated, presum-
ably because of the role of glucose in general metabolism.
The function of yS-galactosidase is to generate glucose by
cleaving lactose, so if both glucose and lactose are avail-
able, the cell can use glucose and there is no reason for
the lac operon to be induced. (The other cleavage product,
galactose, is also converted to glucose by enzymes of the
galactose operon.) Indeed, when glucose is present in the
medium, little or no lac mRNA is made because glucose
TABLE 26-1
Concentration o f Cyclic AM P in Cells Growing in
Media Having the Indicated Carbon Sources
Carbon Source
cAMP Concentration
Glucose
Low
Glycerol
High
Lactose
High
Lactose+ glucose
Low
Lactose+ glycerol
High
indirectly prevents RNA polymerase from binding to the
lac promoter. This positive regulation is accomplished by
cyclic AMP,
a regulatory molecule in both prokaryotes and
eukaryotes. Cyclic AMP (cAMP) is synthesized from ATP
by the enzyme adenylate cyclase, and its concentration is
regulated by glucose metabolism. In a bacterial culture
that is starved of glucose, the intracellular concentration
of cAMP is very high. If a culture is growing in a medium
containing glucose, the cAMP concentration is very low.
The observation that in a medium containing a carbon
source that cannot enter the glycolytic pathway the cAMP
concentration is high (Table 26-1) suggests that some
glucose metabolite or derivative is an inhibitor of adeny-
late cyclase.
Cyclic AMP does not act directly as a regulator but
is bound to a protein called the
cAMP receptor protein
(CRP),
which forms a complex with cAMP (cAMP-CRP).
This complex is active in the lac system and in many
other operons involved in catabolic pathways. The cAMP-
CRP complex binds to a base sequence in the DNA in
the lac promoter region and stimulates transcription of lac
mRNA; thus, cAMP-CRP functions as a positive regula-
tor. The cAMP-CRP complex is not needed for binding of
RNA polymerase to the lac promoter, for a free promoter
binds the enzyme. However, an open-promoter complex
does not form unless cAMP-CRP is also bound to the
appropriate DNA sequence. Thus, when glucose is ab-
sent, the cAMP concentration is high and there is enough
cAMP-CRP to allow an active transcription complex to
form; if glucose is present, cAMP levels are low, no cAMP-
CRP is available, and transcription cannot begin. Thus,
the lac operon is independently regulated both positively
and negatively, a feature common to many carbohydrate
utilization operons (Figure 26-2).
Tryptophan (trp) Operon
The tryptophan operon is responsible for the production
of the amino acid tryptophan, whose synthesis occurs in
