LECTURE 6 - Protein synthesis and structure

Question 1

RNA TOOLS

Answer 1

mRNA : messenger RNA, the RNA that exits the nucleus and bonds with the ribosome

tRNA : transfer RNA, the RNA that brings the amino acids to the ribosomes

rRNA : ribosomal RNA

Question 2

rRNA

Answer 2

Ribosomal ribonucleic acid (rRNA) combines with
proteins to form the machinery for protein
synthesis
AND
catalyses peptide bond formation

Question 3

Amino acyl(aa)-tRNA-synthetases

Answer 3

Aa-tRNA-synthestases
- Are proteins/enzymes
- Attach the correct amino acid to its
matched tRNA
- Recognise the amino acid, the anticodon and other parts of the tRNA
- Catalyse the activation of amino acids
- Use ATP hydrolysis to get the energy to
make a high energy bond`

Question 4

Aa-tRNA-synthetases mechanism

Answer 4

AA is an enzyme
amino acids and ATP binds to this AA enzyme
ATP –> AMP and P-P, (pyrophosphate) is released and P-P slip to form inorganic phosphates
a high energy bond between AMP joins to amino acid
then AMP is replaced by tRNA –> high energy bond between aa and tRNA
–> amino acyl tRNA
amino acyl tRNA is released and is ACTIVATED

Question 5

ribosome

Answer 5

ribosome contains enzymes and riboproteins (made of RNA)

it has E- site : when tRNA exits (Exit)
P- site : growing Protein chain (Peptidyl)
A- site : accepts incoming tRNA-aa (aminoacyl)

Question 6

Inititation fo translation

Answer 6

small subunit of the ribosome
binds mRNA and Met-tRNA (start)
Large subunit of the ribosome
binds = functionin ribosome

Question 7

elongation fo translation

Answer 7

1) AA-tRNA enters the ribosome and comes in guided by anticodon/codon matching; Activated amino acids are positioned next to each other
2) Peptidyl transferase in ribosome catalyses peptide bond between incoming aminco acid and growing polypeptide, using
energy released when the aa-tRNA bond breaks when the amino acid is transferred to growing chain;
3) First tRNA released through the E site of the ribosome, ribosome moves along to next codon on mRNA etc

Question 8

termination fo translation

Answer 8

1) When stop codon is reached,
no tRNA matches; release factor
(which is a protein) binds to stop codon.

2) this signals peptidyl transferase (from
ribosome) to add water, (hydrolysis), releasing the
polypeptide

3) The machinery disassembles,
and the parts can be reused

Question 9

Peptidyl transferase

Answer 9

like the peptidyl links to protein
transferase is like tRNA and ‘ase’ means break down
so tRNA leaves and the amino acids bind together in P site

Question 10

Some challenges of protein synthesis

Answer 10

• The need to convert a sequence of
  nucleotides to a sequence of amino
  acids
  –> need to adapt different
  chemistries
  –> DNA and RNA use bases (A, T, C, G in DNA, and A, U, C, G in RNA), while proteins are made up of amino acids linked together.
• The need to have the correct order of
  amino acids
  –> Any mistake in the sequence can lead to a non-functional or even harmful protein.
  .
• Peptide bond formation is very
  thermodynamically unfavourable

Question 11

addressing challenges of protein synthesis

Answer 11

• The need to have the correct order of
  amino acids
  –> using ribosomes to bring amino acids together
• Peptide bond formation is very
  thermodynamically unfavourable
  –> utilizing enzymes called ribosomes and specific tRNA molecules that assist in the formation of peptide bonds.

Question 12

condensation polymerization to form a dipeptide

Answer 12

This reaction is extremely
thermodynamically unfavourable due
to the large amount of water around.

Hydrolysis (breaking apart/reverse
reaction) is always favoured over
condensation in an aqueous
environment.

Question 13

Prokaryotic versus Eukaryotic Translation

Answer 13

1. Initiation
   Prokaryotic Cells: involves a specialized molecule called the Shine-Dalgarno sequence on the mRNA.

Eukaryotic Cells: rely on a structure known as the 5’ cap and the poly-A tail on the mRNA to find start codon.

different mechanism for finding the start codon
first amino acid is still methionine (Met)

2. Elongation:
   The same in both
3. Termination
   Eukaryotic Cells: a single release factor that recognizes all three stop codons (UAA, UAG, UGA). When a stop codon is encountered, the release factor causes the ribosome to release the completed protein.

Prokaryotic : have two to three different release factors, each specific to one of the three stop codons. When a stop codon is recognized, the corresponding release factor triggers the ribosome to release the protein.

Question 14

antibiotics and prokaryotes translation

Answer 14

many antibiotics target prokaryotic ribosomes
making them effective against bacterial infections without affecting eukaryotic cells

Question 15

primary structure of protein def

Answer 15

• Primary
  The sequence of amino acids bonded by covalent peptide bonds

Question 16

secondary structure of protein def

Answer 16

involves local structures or patterns within the polypeptide chain.
Alpha helix & Beta sheet

Question 17

tertiary structure of protein def

Answer 17

overall three-dimensional arrangement of a single polypeptide chain.
These interactions can include hydrogen bonds, disulfide bonds, hydrophobic interactions, and more.

Question 18

quanternary structure of protein def

Answer 18

Organisation of subunits (Many but not all
proteins have multiple subunits)
when a protein consists of multiple subunits. It describes the organization of these subunits and the interactions between them.

Question 19

Features of secondary structure

Answer 19

1. Secondary structure elements are local features within a protein chain, meaning they involve a short section of the polypeptide. They play a crucial role in determining the protein’s overall three-dimensional structure.
2. Backbone-Backbone Hydrogen Bonding
   –> stabilize the structure
3. Sidechain Interactions and Tertiary Structure
   –> sidechain interactions help hold the secondary structures together.
   –> secondary structures + sidechain interactions + other factors = tertiary structure of a protein –> three-dimensional shape.

Question 20

α-helices + B-pleated sheets (strands)

Answer 20

alpha : Always have same
basic shape, right-
handed helix from N to C
Sidechains
point
outwards

beta : Arrow points
direct of protein
chains (N>C)
Can be parallel (strands
point in the same
directions) or antiparallel
(strands point in opposite
directions)
B-strands packing together to make B pleated sheets
Sidechains
point above
and below

Backbone hydrogen bonding
patterns and bond angles define
the structures

Question 21

haemoglobin and myoglobin

Answer 21

Hemoglobin:
- tetramer, which means it consists of four protein subunits–>come together to form a functional protein complex.
- Each subunit is called a globin, and there are two alpha globin subunits and two beta globin subunits in the hemoglobin tetramer.
- found in RBC and carries oxygen from lungs to body’s tissues/organs

Myoglobin:

monomeric protein, meaning it consists of a single protein subunit.
It does not have quaternary structure because it doesn’t involve multiple subunits coming together.

Myoglobin’s primary function is to store and facilitate the release of oxygen within muscle cells. Its single subunit structure is well-suited for this function.

found in muscle cells, function is to store oxygen in muscle cells

Question 22

the stable conditions for most molecules

Answer 22

• 5 °C to 50 °C
• Atmospheric pressure
• Nearly neutral pH (about 7)

Question 23

α-Helices and B-DNA

Answer 23

• α-helices are a perfect size to fit into the
  major groove of DNA
• Diameter of helix is on average 1.2 nm
• Major groove of B-DNA is 1.2 nm wide
• interacts by binding to DNA for e.g gene regulation, replication, and repair.
• Sidechains must point the right way to
  recognize either the backbone for
  general/non-sequence specific binding, or
  the bases for sequence specific binding
• E.g. positive protein sidechains interact negative
  phosphates in DNA backbone [Positively charged amino acid sidechains on the α-helix can interact with the negatively charged phosphate groups in the DNA backbone.]

Question 24

types of mutations to DNA

Answer 24

1) Silent mutation = no change to protein
sequence
2) No start codon = no protein
3) Frameshift mutation – wrong sequence
4) Single point mutation/missense =single amino acid
change
5) Nonsense mutations introduce stop codon/shorten protein

Question 25

Effect of mutations on Protein
Structure & Function

Answer 25

• Silent mutation – no change (1)
• Frameshift mutation – different protein sequence, protein unlikely to fold correctly (or at all)/may be\ truncated/likely to affect function (2)
• Point mutation =
• Similar residue (e.g. hydrophobic to hydrophobic or similar size) may not have a big effect, minorchange in structure or stability (3)
• Different residue (e.g. hydrophobic to positive charge) may affect folding (especially if inside protein (4)/or function (especially if in binding or active site (5)
• Note that mutations outside of protein coding region can affect gene expression