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DNA
RNA
PROTEIN
FUNCTION
Protein degradation
Protein synthesisAMINO ACIDS
Some properties of intracellular protein degradation (1970).
• Abnormal proteins are rapidly eliminated.
• Normal proteins are selectively degraded at widely different rates.
• Levels of specific proteins in animal cells can be regulated by
changes in rates of synthesis or rates of degradation (Schimke,
1970).
Hershko & Tomkins JBC 1971
Ciechanover, Hod & Hershko, BBRC 1978
APF-1 (ubiquitin)
Protein (µg) 1.5 3 7 PNAS 1980
ATPlysozyme (µg)
- -+ + + + +0 0 5 10 25 5 5
125I-ubiquitin
125I-lysozyme
lysozyme- (Ub)n
Hershko et al., PNAS 1980
PNAS 1980
DUBDUBDUB
E2E2E2 ProteinProtein
E3E3
UbUb
UbUb
UbUb
UbUb UbUb
UbUb
UbUb
UbUb
UbUb
E2E2E2
E1E1E1
UbUb
E1E1E1
+ATP+ATP
UbUb
AMP+PPAMP+PPiiE1E1E1
E2E2E2
UbUb
++ ATPATP
ADP+PADP+Pii
ProteinProteindegradeddegraded
26S proteasome26S proteasome
UbUb
ProteinProtein
UCHUCHUCH
Hershko et al., JBC 1983
Substrate Yes No Yes No Yes Yes
E3α-125I-proteincross-link
125I-protein
Cytochrome c (yeast)
Cytochrome c (mammalian)
Lysozyme
Ribonuclease A (native)
Ribonuclease A (oxidized 1)
Ribonuclease A (oxidized 2)
DETECTION OF E3-SUBSTRATE BINDING BY CHEMICAL CROSS-LINKING
Free 125I-proteinsJBC 1986
MBC 1995
Cyclosome/APC
MBC 1995
UU
UU
UU
UU
Cdk1
Cyclin B
Cdk1
Cyclin B
Cdk1
~ ~~ ~ Cdc20
PP
InactiveCdk1
InactiveAPC/cyclosome
ActiveAPC/cyclosome
E1, E2-CATP, Ub(b)
ATP
(a)
Cyclinsynthesis
26SProteasome
ActiveCdk1
Phosphatase
(d)
Cyclindegraded
Pi
PP
Cdc20
APC/Cinactive
APC/Cactive
APC/Cinactive
APC/C = Anaphase-PromotingComplex/Cyclosome
G0/G1 TimeTime
Lev
elL
evel
S
p27Cyclin E Cyclin A
Skp2
p27Cyclin E
Cdk2
Cyclin E
Cdk2
Cyclin A
Cdk2
Cdk2 inactive Cdk2 activep27 degraded
Cks1
NCB 1999
NCB 1999
The three binding sites of Cks1
Cdk binding(green)Anion binding
(blue)
Skp2 binding(red)
JBC 2002
p27Cyclin E
Cdk2p27
Cyclin E
Cdk2
Cyclin E
Cdk2
Cdk2 inactive
p27-(Ub)n
P
p27Cyclin E
Cdk2
Skp1
SCF complex
P
Skp2 &Cks1
synthesis
Skp2Cks1
Cyclin Esynthesis
Cdk2 active
p27 degraded
Some roles of ubiquitin-mediated protein degradation.
• Control of cell division• Signal transduction• Regulation of gene expression• Responses to inflammation• Immune response• Embryonic development• Apoptosis• Circadian clocks
Some regulatory proteins degraded by the ubiquitin system.
Regulator Role ofdegradation
Examples
I Positive Limitation of duration
Cyclins (G1, S, M-phase); transcription factors (myc, fos…)
II Negative Initiation of process
Cdk inhibitors Anaphase inhibitorIκB transcriptional regulator
III Positive or negative
Activation by stabilization
p53 tumor suppressor; β-catenin
Type
2000
Microbe hunters
Vitaminhunters
Enzyme hunters
Gene hunters
1900
Arthur Kornberg : For the Love of Enzymes (1989)
Technion lab (1971-present)
Dvora GanothHanna HellerEsther EytanSarah EliasClara SegalJudith Hershko
Former graduate studentsAaron CiechanoverYuval ReissValery SudakinShirly Lahavand many others…
Collaboration and helpIrwin A. RoseLeonard CohenJoan RudermanMichele Pagano
Present graduate studentsGil BornsteinDanielle Sitry-ShevahYakir Moshe