2.3.4.1

  • Describe regulation by post-translational proteolysis of proteins.

  • Understand the importance of signaling peptides.

  • Define a zymogen.

  • Work through the processing of chymotrypsin as an example of an enzyme being regulated by post-translational proteolysis.

  • Work through the processing of insulin as a non-enzymatic example of regulation by post-translational proteolysis.


  • Creation of proteins

    • Gene coding for the protein is transcribed into a messenger RNA molecule

    • Second, the messenger RNA molecule (mRNA) is translated into a protein

    • After this the protein will most likely need post-translation modifications and interactions with specific ligands

      • EX) Enzymes depend on the inter with the substrate

      • EX) Enzyme activity is controlled by allosteric modulators

      • EX) Transient covalent medications such as phosphorylation can activate or inactivate some enzymes

    • Synthesized enzymes are sometime made as inactive precursors also known as Zymogens or Proenzymes.

    • These enzymes are activated by triggering a hydrolysis reaction catalysis by a proteolytic enzyme

    • This results usually in a change of conformation of a portion of the polypeptide revealing the active site of the mature fully processed enzyme

    • The processing usually occurs indie the Golgi system, where a specific region of the zymogen is cleaved.

    • These cleaved off regions inactivate the peptide, they are in some cases necessary for the stabilization and proper folding of the zymogen

    • This should not be confused with signal peptides ‘

    • Newly synthesized proteins have different paths they can go on.

      • 1) Some Proteins made on free cytoplasmic ribosomes have no signal sequence and will remain cytoplasmic

      • 2)Other proteins have a specific N-terminal sequence that directs these proteins to mitochondria, nucleus, or other small organelles such as peroxisomes.

      • 3) Some cases, translation is initiated in the cytoplasm, but quickly after the signal sequence has been synthesized, the ribosomes attaches to the surface of the endoplasmic reticulum, translation resumes and proteins accumulate in the lumen of the endoplasmic reticulum this protein will then be transported to the Golgi complex and then secretory vesicles or to a type of cytoplasmic organelles called lysosomes

      • 4) Proteins contained in secretory vesicles get secreted by cells, while those contain in the membrane of these vesicles will be integrated in the membrane of the cell.

  • Signal peptides

    • Located at the N-terminus of proteins

    • Used to direct proteins to specific intracellular compartment

    • Is organelle specific, most cases cleaved off by proteins after devilry

    • Signaling peptides and inactivating peptides represent a powerful mode of regulation by which a protein can be activated where and when needed.

  • Zymogen

    • is a synthesized enzyme

    • Is synthesized as an inactive pre cursor

    • Chymotrypsin

      • Is a digestive enzyme

      • Produced and secreted by the pancreases in an inactive form called chymotrypsin

      • In the duodenum(smallest segment of the small intestine), the active chymotrypsin is a proteolytic enzyme, that breaks down proteins from ingested food.

      • Chymotrypsin is the processed and active form derived from chymotrypsinogen, an inactive zymogen containing 245 amino acids.

        • First step of the activation process is the proteolytic cleavage of the peptide bond between amino acids 15 and 16. This reaction is catalyzed by a protease called trypsin.

        • The end product is the active pi-chymotrypsin, in which two cleaved fragments remain covalently attached via a disulfide bond

        • The final step of the processes, pi-Chymotrypsin takes over and digests itself. Two dipeptide bonds are cleaved off and then released.

        • With the first one containing the residues that occupy position 14 and 15 in pi-chymotrypsin

        • And the second contains residues that occupy positions 147 and 148.

        • The outcome is the mature form of chymotrypsin called alpha-chymotripsin comprised of three polypeptides that remain together because they are linked by sulfide bridges.

  • What does occur at the structural level when chymotrypsinogen

    is converted into chymotrypsin?

    • the side chain of isoleucine, occupying position 16 around the exterior of chymotrypsinogen.

    • Trypsin cleaved the peptide bond between residue 15 and isoleucine 16

    • Upon cleavage, isoleucine 16 can pivot and turn inward. now the free amino acid group carried by isoleucine 16 can form ionic bond with aspartate 194

    • The electrostatic interaction triggers a series of conformational rearrangements, moving methionine 192 from a deeply buried pocket to the surface of the enzyme.

    • The movement of this methionine residue leads to a rearrangement of loops in the surrounding area

    • Induced conformational changes results in the creation of a hydrophobic pocket at the surface of a chymotrypsin.

    • This pocket accommodates the hydrophobic side chains of residues from protein digested chymotrypsin

    • Chymotrypsin cleaves proteins of the C-terminal side of the hydrophobic residues phenylalanine, tryptophan, and tyrosine

    • Proteolytic activation of proteins is not limited to enzymes.

    • Other proteins including hormones and neuropeptides are also activated by proteolysis

  • Insulin Example

    • Insulin is secreted by the pancreases in response to a high blood level of glucose, and stimulates glucose uptake by tissues such as liver adipose tissue, and muscle.

    • Its is transits through the endoplasmic reticulum to the Golgi complex and finally the sector vesicles.

    • Insulin is produced as a 110 amino acid precursor called preproinsulin

      • Its translated at the surface of the endoplasmic reticulum and is pulled through the ER membrane, the N-terminal signal sequence that directs this precursor to the secretion pathway is cleaved off.

      • This results in the accumulation of proinsulin.

        • Proinsulin is then transported to the Golgi complex and to the secretory vesicles

        • Inside the secretory vesicle, insulin is subjected to proteolytic cleavage

        • This results is the production of active insulin formed by the N- and C-Terminal regions that are connected by disulfide bonds

    • Insulin is then formed by two chains called A and B, the region connecting the N- and C-terminal fragments, and produced by proteolysis, is a single polypeptide chain called C-peptide

    • Both mature insulin and the C-peptide are then secreted by the pancreatic cells.

  • C-Peptide

    • Stops the enzyme PDE3 to convert ATP into AMP allowing access ATP to be released into red blood cells, with mature insulin it can adjust the amount of ATP released by the red blood cells and control and rerelease nitric oxide and vasodilation.

    • When white blood cells are activated, they release cytokines that trigger the up-regulation of adhesion molecules at the surface of endothelial cells