Chapter 14: Gene Regulation in Bacteria

Operon

A group of two or more genes under the transcriptional control of a single promoter.

Stem loop

A particular segment of RNA can participate in in the formation of only one of these...? If region 2 forms with region 1, it cannot at the same time form with region 3

Homotetramere

A protein composed of four identical subunits

Lac repressor

A protein that regulates the lac operon by binding to the operator site and repressing transcription; functions as a Homotetramere.

Repressor

A regulatory protein that binds to the DNA and inhibits transcription; negative control

Trans-acting factor

A regulatory protein, such as the lac repressor,

Attenuator sequence

A segment of DNA, important in facilitating this termination

Operator site

A sequence of bases that provides a binding site for a repressor protein called the lac repressor

CAP site

A short DNA segment that functions in Gene regulation; A DNA sequence recognized by an activator protein called Catobolite activator protein (CAP)

Merozygote

A strain of bacteria containing Fʹ factor genes; also called partial diploid.

Promoter

And operon is flanked by a promoter that signals the beginning of transcription

Response to environmental stress

Certain proteins help a bacterium to survive environmental stress such as osmotic shock or heat shock. These proteins are required only when the bacterium is confronted with the stress

Catabolite activator protein (CAP)

Contains 2 subunits, each binding 1 molecule of cAMP; Mediates the effect of cAMP on the lac operon

Lac operon (First unit)

Contains CAP site; lac promoter (lacP); operator site (lacO); three protein encoding genes, lacZ, lacY, lacA; and a terminator.

Presence of both lactose and glucose:

E.coli first uses glucose and Catabolite repression prevents use of lactose

Inducible Genes

Encoded proteins that are produced only when they are required; the cell avoids wasting valuable energy making proteins it does not need

LacA

Encodes galactoside transacetylase, an enzyme that covalently modifies lactose and lactose analogs

LacY

Encodes lactose permease, a membrane protein required for the active transport of lactose into the cytoplasm of the bacterium

Regulatory proteins

Genetic regulatory proteins that respond to small effector molecules typically have two binding sites. One site is where the protein binds to the DNA; the other is the binding site for the effector molecule.

Presence of lactose: glucose used up

If the glucose is used up, catabolite repression is alleviated, and the bacterium then expresses the lac operon. Transcription of lac operon occurs

Effector molecules 1. ⬆️transcription rate 2. ⬇️transcription rate

Inducers: ⬆️ (inducible genes) Corepressors: ⬇️ Inhibitors: ⬇️ (Repressible genes)

Negative control (Inducer genes)

Lac repressor binds to nucleotides within lac operator; this prevents RNA polymerase from transcribing lac Z, lac Y, lac A; in the absence of allolactose(inducer) the lac repressor is bound to the operator most of the time

Presence of glucose:

Leads to the repression of the lac operon

LacI (Second transcriptional unit)

Not part of the lack operon; has it's own promoter called the i-promoter and is constitutively expressed at very low levels. The LAC Lac i gene also encodes the lac repressor

TrpL

Part of lac operon

Stem loops

Region 2 is complementary to regions 1 and 3; region 3 is complementary to regions 2 and 4 3 stem loops: 1-2, 2-3, 3-4

Cyclic-AMP (cAMP)

Small effector molecule which is produced from ATP via an enzyme known as adenylyl cyclase

Allosteric sites

The functioning of allosteric proteins, such as the lac repressor, is controlled by effector molecules that bind to the proteins' allosteric sites.

Diauxic growth

The sequential use of two sugars by a bacterium; is a common phenomenon among many bacterial species.

Catabolite repression

This form of transcriptional regulation is influenced by the presence of glucose, which is a catabolite—a substance that is broken down inside the cell; presence of glucose ultimately leads to repression of the lac operon.

Cis-acting element

a DNA segment that must be adjacent to the gene(s) that it regulates; lac operator site; it is said to have a cis-effect on gene expression.

Trans-effect

a form of genetic regulation that can occur even though two DNA segments are not physically adjacent; the action of the lac repressor on the lac person is considered this; mediated by genes that encode regulatory proteins

Activator

a regulatory protein that increases the rate of transcription; positive control

Inducer

a small effector molecule that causes transcription to increase; 2 ways: bind to a repressor protein and prevent it from binding to the DNA, it could bind to an activator protein and cause it to bind to the DNA (⬆️ transcription)

polycistronic mRNA

an RNA that contains the sequences of two or more genes. Encoded by an operon

Inhibitor

binds to an activator protein and prevents it from binding to the DNA (⬇️ transcription)

Absence of both lactose and glucose (CAP and lac repressor involvement)

cAMP levels are high; binding of the lac repressor inhibits transcription even though CAP is bound to the DNA. Therefore, the transcription rate is very low.

Trp operon components

contains five genes, designated trpE, trpD, trpC, trpB, and trpA, which encode enzymes involved in tryptophan biosynthesis; in addition, two genes, trpR or trpL, play a role in the regulation of the trp operon.

Trp operon

encodes enzymes involved with the synthesis of the amino acid tryptophan; regulated by a repressor protein

Effector Molecule

exerts its effects by binding to a repressor or activator; causes a conformational change in the regulatory protein and thereby influences whether or not the protein can bind to the DNA.

⬆️glucose: ⬇️cAMP

glucose inhibits adenyl cyclase production

TrpR

has its own promoter and is not part of the trp operon; encodes Trp repressor protein

Transcriptional regulation

involves the actions of regulatory proteins that can bind to the DNA and affect the rate of transcription of one or more nearby genes; 2 types of reg proteins: repressor and activator

Gene regulation

is the phenomenon in which the level of gene expression can vary under different conditions.

Attenuation(3)

mRNA from the trp operon is made as a short piece that terminates shortly past the trpL gene; inhibits the further production of tryptophan in the cell.

TrpL and attenuation

mRNA made from the trpL gene contains codons for 14 amino acids that form the trp leader peptide;

2nd key feature of attenuation

mRNA made from the trpL gene has four regions that are complementary to each other, which causes the mRNA to form stem-loops. Different combinations of stem-loops are possible due to interactions among these four regions

Both are present (CAP and lac repressor involvement)

presence of lactose causes the lac repressor to be inactive, which prevents it from binding to the operator site. presence of glucose decreases cAMP levels so that cAMP is released from CAP, which prevents CAP from binding to the CAP site. CAP not bound to CAP site, transcription of lac operon is low in presence of both sugars.

Attenuation(2)

second regulatory mechanism in the trp operon, that is mediated by the region that includes the trpL gene; transcription actually begins, but it is terminated before the entire mRNA is made.

3-4 stem loop uniqueness

stem-loop together with the U-rich attenuator sequence acts as an intrinsic terminator—a ρ-independent terminator; causes RNA polymerase to pause, and the U-rich sequence dissociates from the DNA, and terminates transcription at the U-rich attenuator.

Trp levels ⬇️ in cell

the trp repressor cannot bind to the operator site; RNA Polymerase transcribes; operon is ON

Attenuation(1)

transcription begins, but is stopped prematurely, before most of the trp operon is transcribed.

Only glucose present (CAP and lac repressor involvement)

transcription of the lac operon is very low because the lac repressor is bound to the operator site and CAP is not bound to the CAP site due to low cAMP levels

1st key feature of attenuation

two tryptophan (Trp) codons are found within the mRNA.

Lac I(-) mutations

Result in constitutive expression of lac operon, meaning it is expressed in the presence and absence of lactose; can cause an inability to synthesize any repressor protein; or may produce a repressor protein that is unable to bind to the DNA at the lac operator site.

Cell division

Some proteins are needed for cell division. These are necessary only when the bacterial cell is getting ready to divide

Metabolism

Some proteins function in the metabolism of small molecules. For example, certain enzymes are needed for a bacterium to metabolize particular sugars. These enzymes are required only when the sugars are present in the bacterium's environment

Terminator

Specifies the end of transcription

Allosteric regulation

The action of a small effector molecule, such as allolactose

Only lactose present (CAP and lac repressor involvement)

Allolactose and cAMP levels are high; Allolactose binds the lac repressor, prevents it from binding DNA; cAMP binds CAP, then CAP binds to the CAP site: stimulates the ability of RNA polymerase to begin transcription; proceeds at high rate

Co-repressors

a small molecule that binds to a repressor protein, thereby causing the protein to bind to the DNA (⬇️ transcription)

Trp levels ⬆️

tryptophan acts as a corepressor that binds to the trp repressor protein. This causes a conformational change in the trp repressor that allows it to bind to the trp operator site; operon is OFF

Constitutive Genes

unregulated genes have essentially constant levels of expression in all conditions over time. Unregulated genes are also called; encode proteins that are continuously needed for the survival of the bacterium

LacZ

Encodes the enzyme beta-galactosidase, an enzyme that cleaves lactose into galactose and glucose; beta-galactosidase also converts a small percentage of lactose into allolactose a structurally similar sugar

Enzyme adaptation

Enzyme adaptation refers to the observation that a particular enzyme appears within a living cell only after the cell has been exposed to the substrate for that enzyme.

Lac I(s)

lac operon cannot be induced even in the presence of lactose. These mutations, which are called super-repressor mutations, alter the region of the lac repressor that binds allolactose; results in repressor that can't bind allolactose

Cis effect

mediated by DNA sequences that are binding sites for regulatory proteins