Molecular Basis of Inheritance- 2 Flashcards
Replication, Transcription, Translation
Where does DNA replication occur?
DNA replication occurs in the nucleus in anti-parallel orientation at the “ori” site
Name the enzymes involved in DNA replication
- Helicase
- DNA Gyrase/ Topoisomerase
- SsBP- Single stranded Binding Proteins
- Primase
- DNA-dependent RNA-polymerase
- DNA ligase
Function of Helicase in DNA replication
Helps in separating or unwinding of the double stranded DNA (cutting the wweak hydrogen bonds between corresponding nucleotides).
This creates a replication fork
Function of Gyrase/Topoisomerase
prevents supercoiling of DNA
Function of SsBP
Helps in stabilizing the replication fork (SsDNA)
Function of Primase
At ‘ori’ site
Acts as a recognition site for DNA dependent RNA polymerase with an exposed 3’-OH end
Function of DNA polymerase
RNA dependent DNA polymerase- RNA dependent primer is the recognition site
- Carries out polymerisation and attaches complementary base pairs
- The direction of reading the template by the DNA polymerase is always 3’-5’
- The synthesis of new DNA on the template is always 5’-3’
DNA is antiparallel- reading is in one direction & attaching is in other
Function of DNA ligase
Helps in joining or sealing (ligate) The nicks (gaps) formed during DNA replication
In which strand is synthesis continuous and discontinuous
Continuous in the leading strand
Discontinuous in the lagging strand
Why is polymerization discontinuous in the lagging strand
- Synthesis of the lagging strand has to be discontinuous because DNA polymerase cannot make a new strand in the 3’ to 5’ direction.
- Therefore, a series of Okazaki fragments (short DNA strands), each with their own RNA primer, must be made as to the fork advances.
Differences between leading and lagging strand
- Leading strand: One primer
Lagging strand: Many primers - Leading strand: Continuous polymerisation
Lagging strand: Discountinuous polymerisation
3.
Leading strand: Only one nick is sealed by the DNA ligase
Lagging strand: Several nicks are sealed by the DNA ligase
- Leading strand: No Okazaki fragments formed
Lagging strand: Okazaki fragments formed - Leading strand: 5’-3’ orientation of strand
Lagging strand: 3’-5’ orientation of strand
What is transcription and where does it take place
The process of formation of RNA from DNA is called transcription.
It takes place in the nuclei
In which direction does transcription occur
5’-3’ direction
How is the transcription bubble exposed?
- Before the start of transcription, a short stretch of DNA (~14 bp) gets uncoiled with the help of the enzyme helicase
- The supercoiling is prevented by topoisomerase/gyrase
- The stability of uncoiled SsDNA is maintained by SsBP (Single stranded binding proteins)
- This process exposes a transcription bubble
In which direction are complementary nucleotides being attached on the mRNA
3’-5’ direction
What are exons and introns
Exons: Coding sequences
Introns: Non-coding sequences
What are the non-coding and coding strands?
Non-coding/anti-sense strand: The strand that has the polarity 3’-5’ and acts as a template
Coding/sense strand: The strand that has the polarity 5’-3’ is called the coding strand, even though it doesn’t code for anything i.e. it doesn’t get transcribed at all
What is the logical reason for naming the coding strand this way
The coding strand sequence will be the same as that of the mRNA synthesised (except thymine which is replaced by uracil in the mRNA strand formed)
Name the steps involved in transcription
- Initiation
- Elongation
- Termination
Initiation step of transcription
- RNA polymerase recognises and attaches to a small strip of DNA called the promoter on the template strand.
Elongation step of transcription
- The RNA polymerase moves along the template strand in 3’-5’ direction but adding nucleotides at the exposed 3’-OH end in the 5’-3’ direction
- Synthesis of mRNA occurs in the 5’-3’ direction
Termination step of transcription
- RNA polymerase stops transcription when it reaches the termination site
- It releases itself from the DNA strand and the new mRNA strand moves from the nucleus to the cytoplasm for translation (Eukaryotes)
The final mRNA strand contains both exons (coding sequence) and introns (non-coding sequence)
What are spliceosomes? How do they perform their function?
- The structures that are responsible for removing the introns from the mRNA strand are called spliceosomes.
- They recognise the start and end sequence of introns and cut them and then re-ligate the exons to form a complete functional mRNA consisting of only coding sequences (exons)
- THis sequence is then translocated to the cytoplasm where translation takes place
How is the mRNA modified before getting translocated?
- The 5’ end of mRNA is capped with a modified guanosine triphosphate, and it is called 5’ capping
- Poly A tail (a sequence of adenine residues) is also attached to the 3’ end of mRNA to avoid nuclease attack
What is Lac operon
Lac operon is an operon or a group of genes with a single promoter that encode genes for the transport and metabolism of lactose in E.coli and other bacteria
What is an operon
An operon is a DNA segment containing a few genes that serve to regulate a particular metabolic acitivity. THey are usually expressed as a particular unit (transcribed together)
Name the structural genes IN ORDER + what they code for + function
- lacZ
codes for: β-Galactosidase
function: hydrolyze lactose into glucose and galactose - lacY
codes for: Permease
function: Lactose transporter (into cell) - lacA
codes for: Transacetylase
function: Adds acetyl group to galactose
Lac operon: When lactose is present in the medium
RNA polymerase binds to the Promoter site and the three structural genes are transcribed, Operon is ON
Lac operon: When lactose is absent in the medium
- A repressor protein binds to the Operator site seen between the Promoter site and Structural gene Z (lacZ)
- This prevents the RNA polymerase from moving along the structural genes and they are not transcribed (Operon is OFF)
Lac operon: When both glucose and lactose are present in the medium
- Glucose is metabolized first
- Operon is switched on subsequently for the metabolism of lactose
