Main Ideas - Protein modifications contribute a lot to functional regulation and targeting of proteins and protein pathways!- Phosphorylation is a major regulator involved in almost every signaling pathway in our cells!- O-GlcNac is emerging as a similarily imp. regulator, possibly by counteracting phosphorylation-mediated events!- Acetylation does more than chromatin remodeling-it controls transcriptional pathways, cell cycle, nuclear transport, and possibly most metabolic pathways as well!- Protein glycosylation (if not by GlcNac) functions mostly to target proteins to their appropriate location!- Lipidation of proteins allows them to be anchored into a membrane!- Monoubiquitination and SUMOylation are emerging as major regulators of DNA repair-related processes!!Learning Objectives 1) Recognize the different mechanisms of enzyme regulation. - Protein (enzyme) regulation!- Proteins can be regulated in lots of way! Incl. protein syn, zymogen activation, allosteric regulation, covalent mod, cellular compartimentalization, protein degradation!- Phosphorylation - on Ser, Thr, Try residues!- can be activating or inactivating!- used to reversibly regulate protein function by conformational changes and enable binding events in signal transduction!- regulate many major pathways!- protein kinases: phosphorylate target protein (use ATP)!- some can phosphorylate themselves= autophosphorylation -- can be activating/inactivating!- protein phosphatase: dephosphorylate target protein (release Pi)!- both recog. consensus seq. near target site (ex. SQ/TQ context) !- both can have many diff. targets!- Ex. regulation of translation via phosphorylation!- HRI (heme-reg eIF2 kinase) expressed in RBCs/precursors!- when heme is low, we don't want too much globin so HRI isn't bound by heme --> is an active kinase --> neg. reg. eIF2 (phosphorylates it ) --> eIF2 inactive --> no translation initiation of globin proteins (opp. when heme is high)!- Protein Acetylation - when on histones, increases chromatin accesibility --> increase in txn (normally)!- protein acetyl transferase: acetylates proteins!- can be acetylated!- protein deacetylase: deacetylases proteins!- typ. on a lysine, occur as regulatory events!- reg: chromatin remodeling, txn, cell cycle, nuclear transport, inhibition of phosphorylation!- can be positive or negative reg. depending on the residue!- Protein glycosylation!- Mono-glycosylation: addition of O-GlcNac (on intracellular proteins on Ser or Thr residues)!- Poly-glycosylation: addition of oligosacc chains (glycans) (on membrane proteins or secreted proteins)!- Putting sugar on a protein --> makes it more soluble!- can occur on several aa side chains !- Glycoproteins are made up of: glucose,galactose, mannose, GlcNAc, GalNAc, Fuc, NANA!- Synthesis of O-linked oligosaccs in glycoproteins!- activated monosacc, adding sugar one by one, happens in Golgi!- Synthesis of N-linked oligosaccs in glycoproteins - happens in the ER!- coordinated sugaring of your protein as you make it and transport it into the ER!- put on whole sugar thing at once on Asn residue!- In vivo functions: increases solubility of protein (more H bonds), zip code to target protein to location, protect protein from proteolysis, facilitate secretion of proteins!- Monoglycosylation: O-GlcNAcylation functions in many cell pathways!- txn, translation, cell division!- targets: RNA pol II, nucleoporins, oncogenes, tumor suppressors, cytoskel proteins!- targets same aa's (Ser and THre) as phosphory --> antagonistic events!- Disulfide bond formation - covalent bonds formed between side chains of cysteine residues!- may form spont. or req. help from enzyme (disulfide isomerase)!- can help to form enzyme or get it in a multiprotein complex!- can be on same protein or between 2 diff. proteins!- Covalent mods by lipid as a regulatory event - Lipidation= putting a lipid on your protein!- Protein acylation: covalent mods of aa side chains w/long chain fatty acids. Can occur on:!- Ser of Thr side chain: O-acylation!- Amino terminus: N-acylation!- Cys side chain: Thio (or S-) acylation!- typ. donor= palmitoyl-CoA!- Protein prenylation: addition of isoprenyl units to a protein!- on cystein close to C-term of a protein= CAAX!- typ. donor= pyrophosphorylated lipid!- these mods impart a strongly hydrophobic comp to the protein --> anchor it into a membrane!- Ubiquitination and Sumoylation!- cov mods of a small protein, ubiquitin or SUMO, to a target prot in an isopeptide bond onto a lysine!- they are regulatory and allow for ubi or SUMO-binding prots to form large complexes!- usually activating events!- multiple sumoylation and ubiquitination events reg. a DNA replication protein!!! 2) Understand the general concepts behind various protein modifications in terms of: regulation of activity, location targeting, complex
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