POST TRANSLATIONAL MODIFICATIONS

Describe protein phosphorylation

• target: Ser, Thr, Tyr through OH group (and Pro/Lys when modified with OH)

• target proteins: many

• enzymes: kinase, usually requires ATP (provides phosphate and energy)

• functions: introduces large, bulky, negative side group, alters protein activity

• reversible by: phosphatases

Describe acetylation

• target residue: lysine through side chain amino group

• target protein: histones

• enzymes: HAT (histone acetyltransferase)

• functions:

  • reduces positive charge

  • decreases DNA-histone interactions

  • reduces chromatin condensation

  • increases transcription

• reversible by: HDAC (histone deacetylase) which decreases transcription

Describe hydroxylation

• function: increases H bonding and facilitates phosphorylation by adding OH group

• target residues: proline or lysine

• enzymes:

  • prolyl 3-hydroxylase/prolyl 4-hydroxylase

  • lysyl 5-hydroxyxlase

  • cofactor: vitamin C (ascorbate)

• outcome:

  • hydroxyproline stabilizes collegen triple helix

  • hydroxylysine serves as an attachment site for sugars and stabilizes intra/intermolecular crosslinks

  • scurvy (vitamin c deficiency) results in weakening of collagen because cant obtain OH

Describe carboxylation

• target residue: glutamate (converted to gamma carboxyglutamate)

• target proteins: proteins involved in blood clotting

• enzyme: gamma glutamyl carboxylase which requires vitamin K as a cofactor

  • reduced form of vitamin K (KH2) oxidized to epoxide (KO) in reaction

• function: carboxylation increases negative charges and is required for calcium binding during blood clotting

• applications

  • vitamin k deficiency: greater tendency to bleed

  • warfarin inhibits enzyme that regenerates KH2 and inhibits, therefore, carboxylation reaction

Describe fatty acylation/prenylation

• palmitolyation

  • target: sulfahydryl group of cysteine

  • adds pamitic acid (16 C)

• myristolylation

  • target: glycine of N-terminal, alpha amino group

  • adds acyl group with 14 C

• prenylation

  • target: SH of cys

  • addition of isoprenyl groups to systein near c-terminus via thioether linkage

  • function: membrane targeting and protein-protein interactions during signaling

  • farnesylation (C15: 3×5)

  • geranylgeranylation (C20: 4×5)

Describe glycosylation

• O-linked

  • target residues: S, T, Y, L, P through o-linked glycosidic bond (last two in collogen)

  • 1+ monosaccharides added

  • target proteins: secreted proteins, extracellular matrix proteins

  • subcellular location: o-linked glycosylation occurs in golgi

  • function: molecular recognition

    • most proteins circulating in blood have 0-linked glycosylation

• N-linked

  • target residues: N (amide nitrogen)

  • target proteins: cell surface glycoproteins

  • core-glycosylation in ER

  • complex glycosylation occurs in golgi

  • functions: receptor-ligand interaction

Describe Ubiquitination

• N-terminal Met, 7 lysine residues, C-terminal Gly within ubiquitin

  • monoubiquitination

    • carboxylic acid of C-terminal glycine of ubiquitin forms an isopeptide bond with epsilon amino group of a lysine residue on a target protein

  • polyubiquitination

    • carboxylic acid of c-terminal glycine of a fresh ubiquitin molecule forms an isopeptide bond with epsilon amino group of one of 7 lysine residues or N-terminal amino group of bound ubiquitin

    • chain usually 4+

    • UPS > ubiquitin proteasome system

      • lysine 48 linked polyubiquitin chain targets proteins for degradation through a cytostolic protein degrading machine known as proteasome

    • putting 1 ubiquitin on a target protein involves 3 enzymes

      • e1 - ubiquitin activating enzymes (forms enzyme-ubiquitin complex)

      • e2 - ubiquitin conjugating enzymes (replaces E1)

      • e3 - ubiquitin ligases (passes ubiquitin onto target protien; good drug target)