Lodish H. - Molecular Cell Biology (5ed, Freeman, 2003) (794361), страница 30
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Molecular chaperones arethought to bind all nascent polypeptide chains as they arebeing synthesized on ribosomes. In bacteria, 85 percent of theproteins are released from their chaperones and proceed tofold normally; an even higher percentage of proteins in eukaryotes follow this pathway.The proper folding of a large variety of newly synthesizedor translocated proteins also requires the assistance of chaperonins. These huge cylindrical macromolecular assemblies areformed from two rings of oligomers.
The eukaryotic chaperonin TriC consists of eight subunits per ring. In the bacterial,mitochondrial, and chloroplast chaperonin, known as GroEL,each ring contains seven identical subunits (Figure 3-11b). TheGroEL folding mechanism, which is better understood thanTriC-mediated folding, serves as a general model (Figure3-11a, bottom). In bacteria, a partly folded or misfoldedpolypeptide is inserted into the cavity of GroEL, where it bindsto the inner wall and folds into its native conformation. In anATP-dependent step, GroEL undergoes a conformationalchange and releases the folded protein, a process assisted by aco-chaperonin, GroES, which caps the ends of GroEL.Many Proteins Undergo Chemical Modificationof Amino Acid ResiduesNearly every protein in a cell is chemically modified after itssynthesis on a ribosome.
Such modifications, which mayalter the activity, life span, or cellular location of proteins,entail the linkage of a chemical group to the free –NH2 or–COOH group at either end of a protein or to a reactive sidechain group in an internal residue. Although cells use the 20amino acids shown in Figure 2-13 to synthesize proteins,analysis of cellular proteins reveals that they contain upwardof 100 different amino acids.
Chemical modifications aftersynthesis account for this difference.Acetylation, the addition of an acetyl group (CH3CO) tothe amino group of the N-terminal residue, is the most common form of chemical modification, affecting an estimated80 percent of all proteins:RONCCHHOCH3COAcetyl lysineCH3CNCH2CH2CH2COOCHCH2NH3OPhosphoserine−OPCH2OCHCOONH3O−OH3-HydroxyprolineH2CCHH2CCHHC3-MethylhistidineH 3CNCH2CHCOONH3NOOC -CarboxyglutamateCHNH2CCHCOOOOCCH2CHCOONH3▲ FIGURE 3-12 Common modifications of internal aminoacid residues found in proteins. These modified residues andnumerous others are formed by addition of various chemicalgroups (red) to the amino acid side chains after synthesis of apolypeptide chain.Acetyl groups and a variety of other chemical groups canalso be added to specific internal residues in proteins (Figure 3-12).
An important modification is the phosphorylationof serine, threonine, tyrosine, and histidine residues. We willencounter numerous examples of proteins whose activity isregulated by reversible phosphorylation and dephosphorylation. The side chains of asparagine, serine, and threonineare sites for glycosylation, the attachment of linear andbranched carbohydrate chains. Many secreted proteins andmembrane proteins contain glycosylated residues; the synthesis of such proteins is described in Chapters 16 and 17.Other post-translational modifications found in selected proteins include the hydroxylation of proline and lysine residuesin collagen, the methylation of histidine residues in membrane receptors, and the -carboxylation of glutamate inprothrombin, an essential blood-clotting factor. A specialmodification, discussed shortly, marks cytosolic proteins fordegradation.Acetylated N-terminusThis modification may play an important role in controllingthe life span of proteins within cells because nonacetylatedproteins are rapidly degraded by intracellular proteases.Residues at or near the termini of some membrane proteins arechemically modified by the addition of long lipidlike groups.The attachment of these hydrophobic “tails,” which functionto anchor proteins to the lipid bilayer, constitutes one way thatcells localize certain proteins to membranes (Chapter 5).Peptide Segments of Some Proteins Are RemovedAfter SynthesisAfter their synthesis, some proteins undergo irreversiblechanges that do not entail changes in individual amino acidresidues.
This type of post-translational alteration is sometimes called processing. The most common form is enzymaticcleavage of a backbone peptide bond by proteases, resultingin the removal of residues from the C- or N-terminus of a713.2 • Folding, Modification, and Degradation of Proteins(a)NH2UbAMP+ PPi+ ATPCO E2E1UbE11CCytosolictarget proteinOUb23E3E2OE1 = Ubiquitin-activating enzymeNHE2 = Ubiquitin-conjugating enzymeCUbE3 = Ubiquitin ligaseUb = UbiquitinSteps 1, 2, 3(n times)(b)UbUbUbnCapATP4ADPCoreUbiquitin Marks Cytosolic Proteinsfor Degradation in ProteasomesIn addition to chemical modifications and processing, the activity of a cellular protein depends on the amount present,which reflects the balance between its rate of synthesis andrate of degradation in the cell.
The numerous ways that cellsregulate protein synthesis are discussed in later chapters. Inthis section, we examine protein degradation, focusing onthe major pathways for degrading cytosolic proteins.The life span of intracellular proteins varies from as shortas a few minutes for mitotic cyclins, which help regulate passage through mitosis, to as long as the age of an organism forproteins in the lens of the eye. Eukaryotic cells have severalintracellular proteolytic pathways for degrading misfolded ordenatured proteins, normal proteins whose concentrationmust be decreased, and extracellular proteins taken up by thecell.
One major intracellular pathway is degradation by enzymes within lysosomes, membrane-limited organelles whoseacidic interior is filled with hydrolytic enzymes. Lysosomaldegradation is directed primarily toward extracellular proteins taken up by the cell and aged or defective organelles ofthe cell (see Figure 5-20).Distinct from the lysosomal pathway are cytosolic mechanisms for degrading proteins. Chief among these mechanismsis a pathway that includes the chemical modification of a lysine side chain by the addition of ubiquitin, a 76-residuepolypeptide, followed by degradation of the ubiquitin-taggedprotein by a specialized proteolytic machine. Ubiquitinationis a three-step process (Figure 3-13a):■ Activation of ubiquitin-activating enzyme (E1) by theaddition of a ubitiquin molecule, a reaction that requiresATPTransfer of this ubiquitin molecule to a cysteine residuein ubiquitin-conjugating enzyme (E2)■ProteasomeUbCapUb5UbPeptides▲ FIGURE 3-13 Ubiquitin-mediated proteolyticpathway.
(a) Enzyme E1 is activated by attachment of aubiquitin (Ub) molecule (step 1 ) and then transfers this Ubmolecule to E2 (step 2 ). Ubiquitin ligase (E3) transfers thebound Ub molecule on E2 to the side-chain —NH2 of a lysineresidue in a target protein (step 3 ). Additional Ub moleculesare added to the target protein by repeating steps 1 – 3 ,forming a polyubiquitin chain that directs the tagged proteinto a proteasome (step 4 ). Within this large complex, theprotein is cleaved into numerous small peptide fragments(step 5 ).
(b) Computer-generated image reveals that aproteasome has a cylindrical structure with a cap at each endof a core region. Proteolysis of ubiquitin-tagged proteinsoccurs along the inner wall of the core. [Part (b) fromW. Baumeister et al., 1998, Cell 92:357; courtesy of W. Baumeister.]Formation of a peptide bond between the ubiquitinmolecule bound to E2 and a lysine residue in the targetprotein, a reaction catalyzed by ubiquitin ligase (E3)■This process is repeated many times, with each subsequentubiquitin molecule being added to the preceding one.
The resulting polyubiquitin chain is recognized by a proteasome,another of the cell’s molecular machines (Figure 3-13b). Thenumerous proteasomes dispersed throughout the cell cytosolproteolytically cleave ubiquitin-tagged proteins in an ATPdependent process that yields short (7- to 8-residue) peptidesand intact ubiquitin molecules.MEDIA CONNECTIONSE2E1Overview Animation: Life Cycle of a Proteinpolypeptide chain. Proteolytic cleavage is a common mechanism for activating enzymes that function in blood coagulation, digestion, and programmed cell death (Chapter 22).Proteolysis also generates active peptide hormones, such asEGF and insulin, from larger precursor polypeptides.An unusual and rare type of processing, termed proteinself-splicing, takes place in bacteria and some eukaryotes.This process is analogous to editing film: an internal segmentof a polypeptide is removed and the ends of the polypeptideare rejoined.
Unlike proteolytic processing, protein selfsplicing is an autocatalytic process, which proceeds by itselfwithout the participation of enzymes. The excised peptideappears to eliminate itself from the protein by a mechanismsimilar to that used in the processing of some RNA molecules (Chapter 12). In vertebrate cells, the processing of someproteins includes self-cleavage, but the subsequent ligationstep is absent. One such protein is Hedgehog, a membranebound signaling molecule that is critical to a number of developmental processes (Chapter 15).72CHAPTER 3 • Protein Structure and FunctionCellular proteins degraded by the ubiquitin-mediatedpathway fall into one of two general categories: (1) native cytosolic proteins whose life spans are tightly controlled and(2) proteins that become misfolded in the course of their synthesis in the endoplasmic reticulum (ER).
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