15.12
Modificazioni proteiche nel RER
La modifica delle proteine secretorie e transmembrana che entrano nel RE ruvido inizia nel lume del RE. Queste modifiche aiutano nel ripiegamento dell…
La glicosilazione legata all'N e la formazione di legami disolfuro sono due importanti modificazioni proteiche che si verificano nell'ER.
Durante la glicosilazione, il complesso oligosaccariltransferasi aggiunge molecole oligosaccaridiche ramificate ad alcuni residui selezionati di asparagina di una catena polipeptidica in arrivo.
Al contrario, la formazione di legami disolfuro avviene tra due residui di cisteina ravvicinati sulla stessa catena polipeptidica o su catene polipeptidiche diverse.
Coinvolge due attori chiave: la proteina disolfuro isomerasi, o PDI, e l'ER ossidoreduttasi I, o Ero1.
La molecola PDI assomiglia a un ferro di cavallo con legami disolfuro nel suo sito attivo.
La PDI ossidata ha una conformazione aperta e lega i polipeptidi dispiegati. In primo luogo, il residuo ridotto di cisteina del polipeptide forma un legame disolfuro con il sito attivo dell'enzima.
Questo intermedio interagisce quindi con un'altra cisteina ravvicinata sulla catena polipeptidica, determinando un legame disolfuro tra i due residui.
Successivamente, la PDI cambia in una conformazione chiusa, rilasciando il polipeptide.
Ero1 riconverte la PDI ridotta al suo stato ossidato, preparandola per un altro ciclo di reazione.
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Q1: What is N-linked glycosylation and why does it occur in the ER?
N-linked glycosylation is the covalent attachment of branched oligosaccharide molecules to asparagine residues on incoming polypeptide chains, catalyzed by the oligosaccharyltransferase complex. This modification improves protein folding kinetics, increases stability by masking cleavage sites, and allows ER chaperones to assess proper protein folding before the protein exits the ER.
Q2: How does protein disulfide isomerase facilitate disulfide bond formation?
Protein disulfide isomerase (PDI) resembles a horseshoe with disulfide bonds in its active site. Oxidized PDI binds unfolded polypeptides and forms a disulfide link between a reduced cysteine on the polypeptide and the enzyme's active site. This intermediate then interacts with another nearby cysteine, creating a disulfide bond between the two residues before PDI releases the polypeptide.
Q3: What role does Ero1 play in maintaining the ER redox environment?
ER oxidoreductase 1 (Ero1) recycles reduced protein disulfide isomerase back to its oxidized state, preparing it for additional rounds of disulfide bond formation. Ero1 utilizes a significant fraction of molecular oxygen available in the cell and generates hydrogen peroxide, maintaining redox homeostasis inside the ER while supporting oxidative protein folding.
Q4: Which amino acid sequences are recognized for N-linked glycosylation?
N-linked glycosylation occurs at asparagine residues within the tripeptide sequences Asn-X-Ser and Asn-X-Thr, where X represents any amino acid except proline. The oligosaccharyltransferase complex recognizes these specific motifs and adds oligosaccharides during both cotranslational and post-translational protein translocation in the ER lumen.
Q5: How does PDI function as both an enzyme and a proofreader for disulfide bonds?
Oxidized PDI catalyzes disulfide bond formation between cysteine residues, while reduced PDI acts as a proofreader by correcting inappropriately paired cysteines through rearranging disulfide linkages. This dual function ensures accurate disulfide bond formation and proper protein folding in the oxidizing ER environment.
Q6: What physical properties of proteins are altered by N-linked glycosylation?
N-linked glycosylation improves the thermodynamic kinetics of protein folding, allowing glycosylated proteins to fold better than their non-glycosylated counterparts. Glycosylation also increases protein stability by masking hydrophobic stretches and cleavage sites, protecting the protein from degradation and enhancing its structural integrity.
Q7: Where do disulfide bonds predominantly form in the cell?
Disulfide bonds form predominantly in the rough ER lumen, which provides an oxidizing environment favorable for their formation. A small fraction of disulfide bonds can also form in the mitochondrial intermembrane space, but the ER remains the primary site for this modification of secretory and transmembrane proteins.