Engineering Proteins

EP2 Protein Synthesis

Amino Acids

Some of the 20 Naturally Occurring Amino Acids

Amino Acids - Summary

• Zwitterions• Acid / Base Chemistry (Buffers)• Optical Isomerism – chiral centre, CORN• Peptide – Secondary Amides• Naming dipeptides – NH2 on left• Hydrolysis of peptides• Condensation to form polypeptides and

proteins

Dipeptides

Condensation

Protein Structure

4 levels of structure:-oPrimary Structure – Sequence of Amino Acids;

oSecondary Structure – Spatial arrangement of sections of primary Structure (e.g. helices);

oTertiary Structure – Overall 3D Shape of protein;

oQuaternary Structure – Protein “Monomers” coordinated into tetramers (haemoglobin), or hexamers (insulin).

Primary Structure

Secondary Structure

- Helix - Sheet

Tertiary Structure – gyrase

Quaternary Structure – Insulin Hexamer

Insulin Hexamer

Building Proteins

Cells build proteins directly from L-Amino Acids.

To synthesise a protein, a Chemist would need:-

1) Instructions / knowledge of the primary structure;2) Supplies of pure amino acids;3) A method of forming peptide links.

Cells adopt a similar approach – Protein Synthesis.

Protein synthesis – The role of RNA

1) Messenger RNA - Temporary set of instructions for one protein molecule;

2) Transfer RNA – collects amino acids;

3) Ribosomal RNA – present in ribosomes, which catalyse the formation of peptide links between amino acids.

1) Messenger RNA (mRNA)

2) Transfer RNA (tRNA)

3) Ribosomal RNA (rRNA)

What is RNA? – ribonucleic acid

Nucleic Acids

mRNA triplet base codes (ppt)

RNA Base Pairs

A Permanent set of Instructions

DNA

DNA – deoxyribonucleic acid

1) Codes for many mRNA molecules.

2) The section of DNA coding for a particular protein is called a gene.

3) Full set of genes = genome (humans 3.5 x 109 bases)

Differences between RNA and DNA

DNA Base Pairs

DNA

DNA

How Cells Make Proteins

DNA mRNA

Transcription

Transcription

Transcription – in the nucleus

1) DNA double helix unwinds;

2) hydrogen bonds break;

3) free nucleotides hydrogen bond to the complementary exposed bases;

4) Enzyme (RNA polymerase) links the hydrogen bonded nucleotides to form a strand of mRNA;

5) mRNA is released and the DNA double helix is reformed.

How Cells Make Proteins

DNA mRNA

Protein

Chain

Transcription

Translation

a.a. activation

Amino Acid Activation

Amino Acid Activation

1. tRNA molecule forms an ester link with a specific amino acid;

2. tRNA – amino acid complex moves to the Ribosome.

Translation (ppt)

Translation - in the cytoplasm

1) mRNA attaches to ribosome;2) Hydrogen bonding between complementary bases

binds the correct tRNA anticodon to the 1st codon (set of 3 bases on mRNA);

3) A 2nd tRNA – amino acid complex binds to the adjacent mRNA codon in the ribosome;

4) Ribosome catalyses the formation of the peptide bond between amino acids;

5) The tRNA is released once the amino acid is delivered;

6) Ribosome moves along the mRNA chain to the end.

How Cells Make Proteins

DNA mRNA

Protein

Chain

3D Protei

n

Transcription

Translation

a.a. activation

Folding

Protein Synthesis

How Cells Make Proteins

DNA mRNA

Protein

Chain

3D Protei

n

Transcription

Translation

a.a. activation

Folding