Topic N: Organisation and Control of Prokaryotic Genome

What occurs during specialised transduction?

• Phage: Lambda phage.

• The lambda phage injects the double stranded phage DNA into the bacterial host cell.

• In the bacterial host cell, the phage DNA circularises, and is integrated into the host cell DNA via integrase.

• Thereafter, there is an error in the excision of the phage DNA. As such, the phage DNA is incorrectly excised to include the adjacent regions of the bacterial DNA too.

• The bacterial-phage hybrid DNA is then packaged into the phage capsid during assembly.

• During cell lysis, the phage containing the hybrid DNA then injects another bacterial cell with the hybrid DNA.

• Hybrid DNA will then be incorporated into the bacterial DNA via homologous recombination by replacing homologous regions of the bacterial DNA. This gives rise to genetic variation.

What occurs during generalised transduction?

• Phage: T4 Phage

• The T4 phage injects its double stranded phage DNA into the bacterial cell, and degrades the bacterial DNA into small fragments.

• However, during assembly, fragments of the bacterial DNA are incorrectly packaged into the capsid of a phage.

• Thus, after cell lysis, the bacterial DNA will then be injected into another bacterial cell.

• The foreign bacterial DNA will replace homologous regions of the host bacterial cell’s DNA via homologous recombination. This gives rise to genetic variation.

What are the differences in the transduction processes?

• Specialised transduction occurs during the lysogenic cycle, while generalised transduction occurs during the lytic cycle.

• Specialised transduction primarily involves lambda phage, while generalised transduction often involves T4 phage.

• Specialised transduction occurs due to an error in the excision of phage DNA, while generalised transduction occurs due to an error in the packing of DNA into phage capsid during assembly.

How does conjugation occur?

• The sex pilus from the F+ cell makes contact with the F- cell to form a temporary cytoplasmic mating bridge.

• In the F+ cell, the F plasmid is nicked at the origin of transfer. One strand of the F plasmid is then transferred over to the F- cell via the cytoplasmic mating bridge.

• Now, within both cells, there are single stranded F plasmid. Within each cell, the single stranded F plasmid will be used as a template for the synthesis of a complementary plasmid strand, thus restoring the double stranded plasmid condition.

• This process gives rise to genetic variation.

How does transformation occur?

• The specific cell surface proteins will recognise and transport DNA from closely related species into the bacterial cell.

• The DNA will then replace homologous region of bacteria cell via homologous recombination.

• This process gives rise to genetic variation.

Outline how a prokaryotic genome is organised.

• It is formed by looped domains which are anchored by DNA Binding Proteins, followed by negative supercoiling.

• Found in the nucleoid which is not membrane bound.

• Organised by operons, such hat genes encoding for products involved in the same pathway are controlled by a single promoter.

• As compared to eukaryotic genome, it has fewer genes present, thus making it smaller.

• Absence of introns.

• Circular, double stranded DNA.

• Only has one origin of replication, in contrast to eukaryotic genomes, which can have many origins of replication.

Outline how gene expression is regulated by the trp operon.

• The trp operon is usually turned on, and is a repressive operon.

• However, in the presence of large concentrations of tryptophan, tryptophan will serve as a co-repressors that will bind to the allosteric site of the trp repressors.

• This will cause the trp repressors to undergo a three dimensional conformational change that changes it from the inactive form to the active form.

• The active trp repressors will the bind to the operator, which will prevent the RNA polymerase from binding to the promoter and thus inhibiting the transcription of structural genes downstream.

Outline how gene expression is controlled by the lac operon.

• The lac operon is usually switched off (inducible operon).

• However, in the presence of large concentrations of lactose and low concentrations of glucose, lactose permease will transport lactose into the cell, where the b-galactosidase enzyme will then convert it into allolactose.

• Allolactose will bind to the allosteric site of the lac repressors, causing it to undergo a three dimensional conformational change that changes it from the active form to the inactive form.

• Inactive trp repressors is no longer able to bind to the operator, thus RNA polymerase will be able to bind to the promoter region of the DNA and transcribe downstream structural genes of Lac Z, Lac Y and Lac A. Lac Z codes for b-galactosidase, while Lac Y codes for lactose permease. These genes encode for enzymes involved in the breakdown of lactose.

• When there is low glucose concentration, there will be high concentration of cyclic adenosine mono phosphate which will then bind to the CAP to form a cAMP-CAP complex. This complex will bind to the CAP binding site and increase rate of transcription of downstream structural genes, by helping RNA polymerase bind closely to the promoter.

Outline the differences between both operons.

• Usual state: Lac operon is usually switched off (inducible), while the trp operon is usually turned on (repressible).

• Pathways: Lac operon is involved in a catabolic pathway (breakdown of lactose) while trp operon is involved in an anabolic pathway.

• Effector: Lactose in lac operon, tryptophan corepressor in trp operon.

• Type of Regulation: Dual regulation in lac operon, single regulation in trp operon.