D1.2 Protein synthesis

Describe transcription(happens in nucleus)

1. RNA polymerase binds to promoter region of gene(template strand), unwinding and separating DNA strands

2. RNA polymerase adds RNA nucleotides and forms mRNA by complementary base pairing

3. mRNA strand detaches from template

4. DNA rewinds

5. mRNA leaves nucleus via nucleus envelope pore

Where is rRNA found?

In large subunit of ribosome

True or false: transcription makes a copy of the sense strand of DNA

False, it makes a copy of the antisense (template) strand

True or false: rna polymerase is involved in translation

False, it’s only involved in transcription

True or false: transcription and translation occur simultaneously in prokaryotes

True as they lack a nuclear membrane to separate DNA from ribosomes

Why are DNA templates reliable for transcription?

Highly stable molecule due to H-bonds betw DNA bases & phosphodiester bonds → genetic code doesn’t spontaneously break

In somatic cells that do not divide, the genetic sequence is conserved due to this stability

Transcription as first stage of gene expression

Not even protein is needed in every cell, therefore, only certain genes r expressed:

• Genes expressed r ‘switched on’→undergo transcription & translation

• Genes not expressed ‘switched off’ do not undergo

What is the structure of a ribosome

Consists of a large subunit and a small subunit(EPA)

Made up of proteins and rRNA

What’s the difference between free ribosomes(cytoplasm) and bound ribosomes(RER)

Bound-make proteins used outside of cell

Free-make proteins used within cell

What do polysomes do?

Maximises efficiency of protein synthesis, allowing a cell to produce multiple copies of a specific protein quickly from just one mRNA strand

1. Describe translation

1. In cytoplasm, mRNA attaches to ribosome’s small subunit

2. Initiator tRNA binds to its amino acid and comes to ribosome’s small subunit

3. tRNA contains anti-codon(complementary to mRNA codon)

4. Large ribosomal subunit then binds to form ribosomal complex

5. Initiator tRNA is located in P site, 2nd lands in A site → amino acids on them bind(peptide bond)

6. Ribosome moves so 1st tRNA is in exit site, 2nd in P site → 1st tRNA leaves(to be used again) but leaves amino acid behind

7. Process repeats until stop codon(tRNA without amino acid binds)

True or false: the large ribosomal subunit binds to form a complex AFTER the initiator tRNA binds to the start codon of mRNA on the small ribosomal subunit

True

Why is the genetic code considered degenerate(or redundant)?

There are 4 bases, 4³=64 possible codons (in triplets) while there are only 20 amino acids.

So multiple codons can code for the same amino acids.

Why is the genetic code considered universal?

The same triplet codes code for the same amino acids in all living things (which is why transfer of genes from one species to another is possible)

Why bond forms between 2 amino acids in translation?

Peptide bond

What completes the formation of an amino acid (polypeptide) chain?

A stop codon

What is an advantage of the degeneracy of the genetic code?

They limit the effect of mutations

What is the start codon code in mRNA?

AUG

How many tRNAs can bind simultaneously on a large subunit?

2

What is a gene mutation?

A change in the sequence of bases in DNA(happens during replication)

What is a point mutation?

When one base in DNA sequence is altered(can results in changed amino acid at this location)

Eg of point mutation: sickle cell anaemia: how it happens

1. Mutation changes Hb^A gene to Hb^S

2. Within haemoglobin gene, GAG codon changes to GTG on coding strand

3. Resulting template codon(CAC) is transcribed into mRNA codon GUG instead of GAG

4. Different amino acid replaces og amino acid

Effects of sickle cell anaemia

Haemoglobin S produced instead of haemoglobin A→ causes distortion in red blood cell shape

This shape leads to:

1. Limited oxygen carrying capacity

2. Blocks capillaries n limits flow of normal rbc

What is silent mutation

Changes DNA codon but not amino acid sequence

What is sense mutation(missense)

Amino acid changes completely, altering protein

What is nonsense mutation

Changes codon to stop codon, leads to shortened protein(nonfunctional)

What is point mutation

When a single nucleotide base in DNA is changed

True of false: silent, missense, nonsense mutations r types of point mutations

True

What is the promoter region in a gene in transcription?

Short sequence of bases NOT transcribed

True or false: the promoter region is the first part of a gene to be transcribed

False: it is not transcribed not classed as part of a gene. It’s a non-coding sequence located NEAR the gene

In which direction is the DNA template strand read, and which direction is mRNA synthesised?

Template strand read from 3’ to 5’

mRNA built from its 5’ to 3’

Same as DNA replication

What does the transcription bubble include? (3)

Template strand

Growing mRNA

RNA polymerase

What is the terminator in transcription?

Sequence of nucleotides that, when transcribed, causes RNA polymerase to detach from DNA(stops transcription)

What do transcription factors do in eukaryotes

Attracts and helps RNA polymerase bind to promoter region

They either activate gene to attract RNA polymerase, or deactivate gene to repel

3 stages of transcription

1. Initiation: RNA polymerase binds near promoter, separating DNA strands

2. Elongation: RNA polymerase moves along template→adds 5’ end of free RNA nucleotide to 3’ end of growing mRNA

3. Termination-RNA polymerase reaches terminator sequence→triggers detachment of RNA polymerase n mRNA strand

What are examples of non-coding sequences

1. Regulatory sequences: involved in gene expression by enhancing or suppressing transcription(eg: promoter)

2. tRNA(carries amino acids) and rRNA(makes up ribosomes)

3. Introns: found within genes of eukaryotes(removed during splicing)

4. Telomerase: regions of repeated nucleotide sequence at the end of chromosome→provides protection during cell division(facilitates binding of RNA primer at end of chromosome)

What is splicing and what carries it out?

When introns are removed from pre-mRNA by a spliceosome, and exons are connected together to form mature mRNA

Tip: exons→expressed, introns→intervene

True or false: splicing occurs in both eukaryotes and prokaryotes

False. Only in eukaryotes as prokaryotic mRNA doesn’t have introns

What are spliceosomes composed of?

nRNA (nuclear rna) and proteins

What happens after splicing?

1. A methylated cap is added to 5’ end to protect against degradation by exonucleases

2. A poly-A tail(long chain of A nucleotides) added to 3’ end for further protection and help mature mRNA exit nucleus

What is alternative splicing?

When spliceosome splices exons together in diff ways to make diff proteins (some exons may be removed)

True or false: one gene can only code for one protein (polypeptide)

False. Alternative splicing results in diff combos of exons, allowing one gene to code for diff proteins

Structure of tRNA

Amino acid binding site

Sugar-phosphate backbone

Hydrogen bonds

Anticodon

In which direction is the mRNA strand translated?

5’ to 3’

Which end is an amino acid always attached to on a tRNA molecule? 5’ or 3’?

3’

Steps of initiation of translation

1. Small subunit attached to 5’ terminal of mRNA

2. Initiator tRNA pairs with AUG(start codon of mRNA) at P site

3. Large subunit of ribosome is added

Function of each ribosome site (APE)

A- holds tRNA carrying next AA being added

P- holds tRNA carrying growing polypeptide chain

E- releases tRNA that lost its AA

What are the sequences of the 3 stop codons?

UAA, UAG, UGA (brought to mRNA by a tRNA molecule that doesn’t contain an amino acid)

State the phases of transcription and translation

1. Initiation

2. Elongation

3. Termination

True or false: tRNA helps translate anticodons into amino acids

False: it translates mRNA codons into amino acids

Describe the modification of polypeptides into their function state. (Include 2-stage modification of pre-pro insulin to insulin)

1. Pre-proinsulin is attached to the endoplasmic reticulum

2. It’s modified by an enzyme which removes the signal peptide from the end→this detaches it from the ER and converts it to proinsulin

3. Proinsulin folds, disulfide bonds form (tertiary protein)

4. Proinsulin is packed into vesicles at Golgi apparatus and its C peptide is removed

5. This results in 2 protein chains attached tg by disulfide bonds→final insulin can be released

Which chemical labels unneeded, damaged or misfolded proteins?

Ubiquitin→it begins the process of these proteins being broken down in the proteasome

What is a proteasome and what does it do?

It’s an organelle in eukaryotic cells that contains protease enzymes that break down peptide bonds of unnecessary/damaged proteins

Amino acids are then released and used in translation to make new proteins

True or false: the number of T nucleotides on one strand of DNA is equal to the number of A nucleotides on the other strand

True, same applies to G and C

What is a proteome

The complete set of proteins express by an organism

Unlike the genome (DNA) that stays the same throughout a lifetime, a proteome constantly changes

True or false: complementary bases in DNA and RNA are held together by covalent bonds

False: they are held by hydrogen bonds

True or false: translation involves both rna and dna

False: it involves only rna

How many sites do both free and bound ribosomes each have and what are they for?

4 sites

3 for tRNA and 1 for mRNA