DEGRADACIJA PROTEINA
Cartoon depiction of cotranslational folding of a polypeptide. The nascent polypeptide is shown assuming secondary
structure as it emerges from the ribosome during the process of biosynthesis. The earliest intermediate, I1, is not wellstabilized by extensive tertiary interactions and is in equilibrium with multiple conformations. The second intermediate
shown, I2, is the N-terminal domain; more extensive tertiary interactions will allow this intermediate to be more stable. The
final intermediate, I3, depicts the structure of the full-length polypeptide immediately prior to release from the ribosome with
the C-terminal domain not yet fully packed. Chaperones and/or folding catalysts (CH and FC) may interact with either the
nascent intermediate structures or with the full-length product (M*) following release from the ribosome. The final stages of
folding from M* to native monomer, Mn, occur following release. Association of monomeric units into oligomeric structures,
O, may occur posttranslationally as depicted or, as discussed in the text, may involve nascent polypeptides. The structure
that was used to develop this cartoon was of one subunit of the bacterial luciferase b2 homodimer (66).
Suvišne komponente u stanici se
proteolitički degradiraju
Dva puta degradacije proteina
A scenario for GHR endocytosis and downregulation: (1) GH binds and dimerizes two GHRs, which causes recruitment
and activation of Jak2; Jak2 phosphorylates the GHR, itself, and STATs. Signal transduction is relayed to the nucleus and
genes are activated; (2) specific ubiquitin conjugases (E2) and ligases (E3) dock onto the UbE motif and ubiquitinate the
GHR and possibly other attached proteins; (3) the E2/E3s activate the clathrin-coated endocytosis machinery, serving either
as adaptors for clathrin or as connectors to AP2 adaptors; (4) transport to early and late endosomes; (5) degradation of GH
and GHR by proteasomes and lysosomes. Journal of Cell Science 112, 1417-1423 (1999)
Ubiquitination motifs
Journal of Cell Science 112, 1417-1423 (1999)
Ubikvitin i proteasomi
Konjugiranje s ubikvitinom
Primjer:
Degradacija ciklina
tijekom staničnog
ciklusa
The ubiquitin system
of S. cerevisiae
The yeast ubiquitin genes, two of which (UBI1 and
UBI2) contain introns, encode fusion proteins of
ubiquitin (yellow rectangles) to itself (UBI4) or to one
of the two specific ribosomal proteins (UBI1–UBI3)
(red and blue rectangles). These fusion proteins are
cleaved by deubiquitinating enzymes, yielding mature
ubiquitin.
Thioester bonds between ubiquitin and the active-site
Cys residues of ubiquitin-specific enzymes.
The conjugation of ubiquitin to other proteins involves
a preliminary ATP-dependent step, in which the last
residue of ubiquitin (Gly76) is joined, through a
thioester bond, to a Cys residue in the ubiquitinactivating (E1) enzyme encoded by UBA1.
The activated ubiquitin is transferred to a Cys residue
in one of at least 13 distinct ubiquitin-conjugating (E2)
enzymes encoded by the UBC family genes, and from
there to a Lys residue of an ultimate acceptor protein
(yellow oval).
This last step and the formation of a multi-ubiquitin
chain (black ovals) require participation of another
component, called E3 (the names of some of the
yeast E3 proteins are included). A targeted,
ubiquitinated protein substrate is processively
degraded to short peptides by the ATP-dependent
26S proteasome.
The N-end rule pathway
N-degron
Notations in the yeast (a) and mouse (b) pathways show
type 1 (purple) and type 2 (red) primary, secondary (light
blue) and tertiary (green) destabilizing N-terminal residues;
yellow ovals indicate the rest of a protein substrate.
a, The in vivo half-lives of X-gals, -galactosidase-based
test proteins in S. cerevisiae (right). X-gal proteins bearing
stabilizing N-terminal residues (black) are metabolically
stable (t1/2, more than 20 h). The tertiary destabilizing
residues N (Asn) and Q (Gln) are converted into secondary
destabilizing residues D (Asp) and E (Glu) by N-terminal
amidohydrolase (Nt-amidase), encoded by NTA1. D and E
are conjugated to R (Arg), one of the primary destabilizing
residues, by Arg–RNA protein transferase (R-transferase),
encoded by ATE1.
b, In the mammalian N-end rule pathway, the deamidation
step is mediated by two distinct enzymes, NtN-amidase
and NtQ-amidase, specific for N-terminal Asn and Gln
residues, respectively54. In vertebrates, the set of
secondary destabilizing residues contains not only Asp
and Glu but also Cys (C), which is a stabilizing residue in
yeast55. In mammals but not in yeast, Ala (A), Ser (S) and
Thr (T) are primary (type 3) destabilizing residues48. c, S.
cerevisiae UBR1 has two binding sites for the primary
destabilizing N-terminal residues of either proteins or short
peptides. The type 1 site is specific for basic N-terminal
residues Arg, Lys and His. The type 2 site is specific for
bulky hydrophobic N-terminal residues Phe, Leu, Trp, Tyr
and Ile. UBR1 contains yet another substrate-binding site
(i), which targets proteins bearing internal (non-N-terminal)
degrons. In yeast, these proteins include the CUP9
repressor57. A complex of UBR1 and the ubiquitinconjugating (E2) enzyme RAD6 produces a substratelinked multi-ubiquitin chain
Protasomi
20S Proteasome
The catalytic core of the proteasome (20S proteasome)
is built up of seven different a and seven different bsubunits arranged as a cylindrical 777  7 complex in
four stacked rings.
Vertebrates possess three additional -subunits that are
induced by interferon- (IFN-).
Degradation of folded proteins requires cooperation of
the 20S-core protease with the 19S cap, a structure that
can bind to both ends of the barrel, resulting in the 26S
proteasome.
The 19S cap contains at least 20 subunits, some of
which are involved in the recognition and removal of
ubiquitin. It also possesses an ATPase activity that helps
to unfold and feed the substrate into a chamber in the
20S core that harbours the active sites.
Further complexity arises from an additional 11S
regulatory complex, also upregulated upon stimulation
with IFN-. Two forms of the 11S regulator exist: a
heterooligomeric ring made up of REGa and REGb
subunits and a homo-oligomeric REGg particle. The 11S
regulator can replace the 19S complex and results in
activation and altered peptidase activity upon binding
to the 20S proteasome in vitro.
Intracellular targeting of proteasome
Trends in Cell Biology,10:268, 2000.
Multiubiquitinated membrane proteins
target the proteasome to the
endoplasmic reticulum (ER).
Ubiquitin-activating enzyme E1
hydrolyses ATP and forms a high-energy
thioester linkage between a cysteine of
its active site and the C-terminus of
ubiquitin. Activated ubiquitin is then
transferred to a ubiquitin-conjugating
enzyme E2. These enzymes function in
concert with ubiquitin protein ligases, E3,
and attach ubiquitin to the substrate
protein. Isopeptidases associated with
the 19S cap or in the cytosol
disassemble the multiubiquitin chain and
ubiquitin can be recycled.
Proteasomi za antigensku prezentaciju
Perfect use of imperfection
Nature 404, 709 - 710 (2000)
Figure 1 A use for protein waste. A significant amount (30% or more) of protein production is faulty and results in the formation
of defective ribosomal products (DRiPs). Cellular quality-control mechanisms ensure that DRiPs are immediately removed by
becoming tagged with a small molecule called ubiquitin (Ub) and targeted to the proteasome, which chops the DRiPs up into
small peptides. Samples of the peptides are transported by the transporter associated with antigen presentation (TAP) into the
endoplasmic reticulum (ER), loaded on MHC class I molecules and transported to the cell surface for recognition by T cells.
Peptides from fully functional proteins that have aged are presented to T cells in the same way. But the DRiP short cut ensures
that the immune system can get a head start on fighting infection, because it does not have to wait for proteins (such as those
produced in virus-infected cells) to age.
Dvije glavne obitelji molekularnih chaperona
Chaperonin
Djelovanje Hsp70 i Hsp60