Bacteria

Question 1

Bacteria

Answer 1

Prokaryotes with no membrane-bound organelles

Question 2

Parts of Bacteria

Answer 2

1. Internal structure
2. Surface structure
3. Appendages

Question 3

1. Internal structure (5)

Answer 3

1. Bacterial chromosome
   - Double-stranded, circular DNA
   - Within nucleoid region, no membrane
   - Associates with DNA binding proteins → loop domains → supercoiling
   - No introns
2. Nucleoid
   - Region in bacteria cell where DNA confined to
3. Ribosomes
   - 70S → granular appearance
4. Storage granules
5. Plasmid
   - Small, circular autonomously replicating DNA molecule
   - Confer advantages
   - Can be more than 1 copy
   - Can have different plasmids
   - Vectors in genetic engineering

Question 4

2. Surface structure (3)

Answer 4

1. Cell membrane
   - Phospholipid bilayer
   - ETC and ATP synthase
2. Cell wall
   - Peptidoglycan → long chains of sugar cross-linked by short peptide chains
   - Protects cell from osmotic lysis
   - Confers rigidity and shape
   - Gram stain
3. Capsule
   - Glycocalyx (sugar coat) → exterior of cell wall
   - Capsule or slime layer
   - Protects bacteria from phagocytosis as it can’t be recognised
   - Enables bacteria to adhere to one another or particular surfaces → biofilm
   - Prevent desiccation

Question 5

3. Appendages

Answer 5

1. Fimbriae (Fimbria)
   - Short, bristle-like fibres
   - Attachment to surfaces/other bacteria/organisms
2. Pili (Pilus)
   - Longer and fewer in number than 1
   - Conjugation → sex pilus
   - Motility → makes contact with surface and retracts to pull
3. Flagella
   - Long appendages for motility
   - Hollow cylindrical protein thread → propels by rotation

Question 6

Binary Fission

Answer 6

Asexual reproduction produces genetically identical bacteria

1. DNA replication begins at the origin of replication (ori) → DNA unzipped by breaking H bonds between bases of the 2 strands to form replication bubble
2. DNA replicates by semi-conservative replication → each original strand serves as template for synthesis of daughter strands by cbp
3. 2 newly formed ori move to opposite poles of the cell and attach to the plasma membrane
4. Cell elongates to prepare for division.
5. DNA is circular with no free ends → 2 daughter DNA molecules will be interlocked with completion of replication.
6. Enzyme topoisomerase cuts, separates and reseals the 2 DNA molecules
7. Invagination of plasma membrane and the deposition of new cell wall (division septum) eventually divides parent cell into two daughter cells → each inherits a complete genome (genetically identical)

Question 7

Genetic variation

Answer 7

Generate genetic variation through forming new combination of new alleles → enhancing reproductive success

Question 8

Genetic variation methods (3)

Answer 8

1. Transformation
2. Transduction (General/Specialised)
3. Conjugation

Question 9

Transformation definition

Answer 9

Uptake of naked, foreign DNA from the surrounding environment, resulting in a change of the bacterial cell’s genotype and phenotype

Question 10

Transformation (7)

Answer 10

1. Fragments of foreign naked DNA from dead lysed bacterial cells
2. Naturally competent bacteria with cell-surface proteins bind and transport DNA into the cell.
3. Artificially, bacteria can be made competent through immersion in a medium with CaCl2 followed by a heat shock treatment
4. Foreign DNA incorporated into chromosome through crossing over at 2 homologous regions found on the bacterial chromosome (homologous recombination)
5. Result: recombinant cell
6. If different alleles for a gene were exchanged, the new allele will be expressed → permanent change in genotype & phenotype
7. Recombinant genome will be passed on to all subsequent offspring through binary fission

Question 11

Transduction definition

Answer 11

Bacterial DNA from one host cell is introduced into another bacterial host cell by a bacteriophage due to aberrations in the phage reproductive cycle

Question 12

Generalised Transduction (5)

Answer 12

1. A phage infects a bacterium, injecting its viral genome (DNA) into the host cell
2. Bacterial DNA degraded into small fragments, one of which may be randomly packaged into a capsid head during the spontaneous assembly of new viruses
3. Upon cell lysis, the defective phage will infect another bacterium and inject bacterial DNA from the previous host cell into the new bacterium
4. Foreign bacterial DNA can replace homologous region in the recipient cell’s chromosome if crossing over/homologous recombination* takes place, possibly allowing expression of a different allele from previous host.

Question 13

Specialised Transduction (6)

Answer 13

1. Temperate phage infects bacterium, injecting viral genome into host cell
2. The viral DNA is integrated into bacterial chromosome forming a prophage which may be improperly excised to include adjacent segment of bacterial DNA during an induction event
3. Bacterial DNA may be packaged into a capsid head during the spontaneous assembly of new viruses
4. Upon cell lysis, defective phage will infect another bacterium and inject bacterial DNA from previous host cell into new bacterium
5. Foreign bacterial DNA can replace homologous region in the recipient cell’s chromosome if crossing over/homologous recombination take place, possibly allowing expression of a different allele from previous host

Question 14

Conjugation definition

Answer 14

Direct transfer of genetic material from one bacterial cell to another through a mating bridge between the two cells via the transfer of F plasmid from an F+ donor to F– recipient cell

Question 15

Conjugation (5)

Answer 15

1. Sex pilus (coded for by F factor) of F+ bacterial cell makes contact with a F- cell and retracts to bring the 2 cells closer
2. The hollow pilus then acts as a cytoplasmic mating bridge between the 2 cells
3. One of the 2 strands of the plasmid DNA is nicked and transferred from the F+ cell to the F- cell through the bridge
4. The single stranded F plasmid DNA circularises in F - cell and is used as a template to synthesise a complementary strand for a double-stranded
   plasmid DNA. The F- recipient cell is now a F+ cell
5. Replication of the plasmid occurs via rolling circle DNA replication occurs

Question 16

Rolling circle DNA replication (5)

Answer 16

1. 1 strand of ds F plasmid nicked by nuclease
2. Free 3’OH end then used as a primer for strand elongation by DNA polymerase using intact strand as a template
3. Elongation process is facilitated by the displacement of the 5’ end of the nicked strand and is transferred across the mating bridge to the recipient bacterium
4. Upon completion of a unit length of the plasmid DNA (after 1 round), another nick occurs to release the
   original strand
5. In the recipient cell, the single strand of F plasmid DNA re-circularises and serves as a template for the synthesis of a complementary daughter stand to form a double stranded circular DNA.

Question 17

Gene Regulation

Answer 17

• Rate at which certain protein molecules are synthesised varies according to circumstances and demand
• Predominates at transcriptional level → efficient with minimal wastage
• Economical use of energy and resources
• Responsiveness to environment
• Selective advantage

Question 18

Operon

Answer 18

A cluster of genes with related functions, regulated in such a way that all the genes in the cluster are turned on and off together

1. Common promoter
2. Operator
3. ≥ 1 structural genes → controlled as a unit to produce a single polycistronic mRNA

Question 19

Promoter

Answer 19

RNA polymerase binding site, upstream of structural genes

Question 20

Operator

Answer 20

Repressor protein binding site to prevent RNA polymerase from binding to the promoter and intiating transcription

Question 21

Polycistronic mRNA

Answer 21

• mRNA that contains base sequence coding for the amino acids sequence of several proteins.
• Contains multiple start and stop codons (one per polypeptide)
• Gives rise to a total of ___ different polypeptides which can be translated from a single mRNA

Question 22

Structural gene

Answer 22

Any gene that codes for a protein product that forms part of a structure or has an enzymatic function

Question 23

Regulator gene

Answer 23

• Any of several kinds of nucleotide sequences involved in the control of the expression of structural genes
• Codes for a protein involved in regulating the expression of other genes e.g. repressor, CAP
• Has its own promoter and terminator sequences
• Not within operon, usually far away, but gene products that control the expression are diffusible

Question 24

Effector

Answer 24

• Small molecule that binds to a specific protein, causing a conformational change and hence regulating its biological activity.
• In this context, includes inducer (allolactose in lac operon) and corepressor (tryptophan in trp operon)

Question 25

Lac operon

Answer 25

- E, coli → intestines of humans and mammals - Inducible operon - Catabolic pathway - Breakdown of lactose - Dual control → negative/positive regulation

Question 26

Lac operon structure

Answer 26

- LacZ, lacY, lacA - Promoter → RNA pol bonding site - Operator → lac repressor binding site - Operator overlaps with promoter - Catabolite Activator Protein (CAP) binding site within promoter

Question 27

LacZ codes for:

Answer 27

β-galactosidase

Question 28

β-galactosidase function (2)

Answer 28

1. Hydrolyse lactose into glucose and galactose | 2. Convert lactose into allolactose, an isomer

Question 29

LacY codes for:

Answer 29

Permease, a membrane transport protein

Question 30

Permease function

Answer 30

Facilitates movement of lactose from outside the cell to inside

Question 31

LacA codes for:

Answer 31

Transacetylase → function not very well known, metabolises certain disaccharides

Question 32

Lac operon regulatory gene

Answer 32

LacI that codes for lac repressor protein

Question 33

Lac operon effector

Answer 33

Inducer lactose, substrate

Question 34

Lac operon default (4)

Answer 34

1. Repressed by default 2. Regulatory gene lacI constitutively transcribed → lac repressor protein 3. Lac repressor protein produced in active form → binds to lac operator via DNA-binding site → denying RNA polymerase access to promoter 4. Transcription of structural genes blocked

Question 35

Lac operon negative regulation (6)

Answer 35

1. In the absence of lactose, a basal level of beta galactosidase and permease is present in the cell: repression of lac operon by lac repressor is leaky 2. Lactose enters the cell by permease 3. And is converted to allolactose by β-galactosidase 4. Allolactose acts as an inducer and binds to allosteric site of lac repressor → change in confirmation → inactive 5. The inactive repressor can no longer bind to operator of lac operon (no longer complementary in shape and charge) 6. Promoter site available for RNA polymerase to bind

Question 36

Lac operon positive regulation (3)

Answer 36

1. When glucose is absent → high levels of cAMP is present → binds to CAP → activated CAP binds to promoter of lac operon → ↑ affinity of RNA pol to promoter 2. Transcription frequency of structural genes lac Z, lac Y and lac A to produce beta-galactosidase, permease and transacetylase respectively to breakdown lactose thus ↑ 3. (Give one example of a product and what it does)

Question 37

Trp operon

Answer 37

- Repressible operon (end-product repression) - Anabolic pathway - Synthesis of amino acid tryptophan (corepressor) - 5 structural genes (trpA-E) - Regulatory gene → trpR → trp repressor

Question 38

Trp operon default (3)

Answer 38

1. Regulatory gene, trpR, constitutively transcribed → trp repressor protein 2. Trp repressor protein synthesised in inactive form → unable to bind to operator 3. RNA pol able to bind to promoter → operon is default "on"

Question 39

Trp operon negative regulation

Answer 39

1. When tryptophan is present in high concentrations in the cell, it acts as a corepressor and binds to the of trp repressor 2. This causes a change in conformation of trp repressor and trp repressor become active → can bind to the operator (complementary in shape and charge) 3. Prevents binding of RNA polymerase to promoter and transcription of structural genes & expression of operon 4. Synthesis of tryptophan is reduced/stopped