Intracellular proteolysis Flashcards
What are proteases / proteinases / peptidases / cathepsins?
They are all enzymes that cleave peptide bonds in the context of proteins.
Cathepsins - They are lysosomal proteins.
Name some different types of proteases?
- Serine proteases - Has a serine residue that plays a critical role in catalysis at active site.
- Cysteine proteases - Has a cysteine residue ……
- Aspartyl proteases - Has an aspartic residue ……
- Metalloproteases - Any proteases enzyme whose catalytic mechanism involves a metal. E.g. Meltrin has a significant role in myogenesis which is the fusion of muscle cells during embryo development.
How do we classify proteases based on the manner in which they bind to and cleave their targets?
There are 2 types of proteases:-
- Endopeptidases:- The enzyme cleaves its substrate protein somewhere in the middle of the peptide chain..
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Exopeptidases:- cleave off just 1-3 amino acids from the end of the protein. They have 2 further subdivision-
- Aminopeptidases - cleaves peptide bonds at the N - terminal (amine terminus).
- Carboxypeptidases - Cleave peptide bonds at the C - terminal (carboxy terminus).
When does the sequence specificity of proteolysis need to be specific?
When he proteases cleave their substrate for the purpose of protein activation.
E.g in insulin.
When does the sequence specificity of proteolysis need to be non-specific?
When the proteases cleave their substrate for the purpose of protein degradation.
E.g. Lysosomes.
How are digestive enzymes activated by proteolysis?
- These enzymes are generated by the exocrine pancreas and secreted into the small intestine.
- Chymotrypsin and trypsin are very abundant enzymes involved in protein degradation during digestion.
- The precursor of chymotrypsin is called chymotrypsinogen. It is initially a peptide of 245 amino acids and it need to be activated by the action of a different enzyme called trypsin.
- The cleave is formed between amino acids 15 and 16 (between arganine and leucine).
- Chymotrypsin itself in the process of autocatalysis or autolysis cleaves itself again which leads to two different sites which releases two very small peptides (serine-14 and serine-15).
- 3 amino acids are removed and so three peptide chains are made which are joined by disulfide bonds.
What is the cause of X-linked haemophilia?
- Deficiencies of factor VIII or IX are the cause of X-linked Haemophilia.
Haemophilia is an inherited condition that affects the bloods ability to clot.
What are some examples of cysteine proteases?
- Bromelain
- Papain
Both can be used as meat tenderisers.
What is an example of a known aspartyl protease?
HIV-1 protease (retropepsin) which cleaves poly-protein precursors to form functional proteins.
What is ubiquitylation?
It is the addition of the ubiquitin ‘tag’ that targets the enzyme to the proteasome for degradation.
Proteasomes - are protein complexes which degrade unneeded or damaged proteins by proteolysis, a chemical reaction that breaks peptide bonds. Enzymes that help such reactions are called proteases.
What are the three enzymes we need to know involved in ubiquitylation?
- E1: Ubiquitin-activating Enzyme.
- E2: Ubiquitin-conjugating Enzyme.
- E3: Ubiquitin-protein Ligase
Describe the three-step pathway for protein ubiquitylation.
- A thioester bond forms between the COOH terminus of ubiquitin and a cysteine in ubiquitin-activating enzyme (E1). This reaction requires ATP.
- The ubiquitin is transferred from cysteine onto ubiquitin-conjugating enzyme (E2).
- The ubiquitin-protein ligase (E3) transfers ubiquitin from E2 to a lysine on the target protein.
- Many different E3s exist that recognise specific target proteins.
What is the half life of proteins?
Half life is the time taken for a protein to degrade in half and measure how stable the protein is.
What is the N-end rule?
- The N-end rule is a rule that governs the rate of protein degradation through recognition of the N-terminal residue of proteins.
- The rule states that the N-terminal amino acid of a protein determines its half-life (likelihood of being degraded).
What is the SREBP cycle for?
It is for the regulation of cholesterol.
What happens in the SREBP cycle when the cholesterol levels are low?
- Ubiquitin targets Insig for degradation.
- Secretory proteins escort SCAP/SREBP to the Golgi complex.
- In the Golgi, two proteases release regulatory domain of SREBP.
- The regulatory domain enters the nucleus and transcription of lipid synthesising enzymes are simulated.
- This increases the cholesterol synthesis.
