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Pyruvate Kinase M2 : linking glycolysis to amino acid biosynthesis
The presented work was done in collaboration with Astex Pharmaceuticals
I am a consultant of Astex Pharmaceuticals
I will not discuss off label use and/or investigational use in my presentation
Disclosure Information2012 AACR annual meeting
Eyal Gottlieb
PK
LactatePyruvate
PEP
3PG
Glucose
GA3P
Respiration
G6P
E1PKLR Gene1q21
E2 E3 E4 E5 E12
CAAT TATA
PKRmRNA E1 E2 E3E4E5
PKLmRNA E2 E3E4E5 E6-E12
E6-E12
Pyruvate Kinase L/R isoforms
E10
E9 E10E8 E12E11
E9E8 E12E11 E10E8 E12E11
PKM1 mRNA PKM2 mRNA
PKMgene15q22
Pyruvate Kinase M1/M2 isoforms
B Domain
A Domain
C Domain
Active Site
FBP
Tyr105 (phosphorylation)
Lys305 (acetylation)
Cys358 (oxidation)
Pro403/ 408 (hydroxylation)
180º
Inter-subunit interface
PKM2
LactatePyruvate
PEP
3PG
Glucose
GA3P
PPP
Serine
Respiration
Glycine
G6P
ROS
Nucleotides
Phospholipids
Cancer cell lines express high levels of PKM2
Silencing PKM2 expression does not change ATP levels and proliferation capacity of HCT116 cells
Metabolic adaptation to PKM2 silencing in HCT116 cells
PKM2 silencing diverts more glucose-derived carbons to serine/glycine biosynthesis and oxidative
phosphorylation
Metabolic adaptation to PKM2 silencing in HCT116 cells
Serine and glycine starvation decreases PK activity
Serine binds to and activates PKM2 in vitro
PKM2 H464A
[compound] (M)
% a
ctiv
atio
n
10- 1 0 10- 8 10- 6 10- 4 10- 2 100
0
1000
2000
3000
4000SerFBP
PKM2 S437Y
[compound] (M)
% a
ctiv
atio
n
10- 1 0 10- 8 10- 6 10- 4 10- 2 100
0
200
400
600
800SerFBP
ITC data PKM2 WT PKM2 H464A PKM2 S437YKd for Ser 200µM N/B 210µM
Mutation of the amino acid binding pocket abolishes serine activation
Serine specifically activates PKM2but not PKM1 or PKLR
PKLR
[compound] (M)
Fold
-act
ivat
ion
(rel
ativ
e to
no
enzy
me
cont
rol)
10- 1 0 10- 8 10- 6 10- 4 10- 2 1000
5
10
15SerFBP
PKM1
[compound] (M)
Fold
-act
ivat
ion
(rel
ativ
e to
no
enzy
me
cont
rol)
10- 1 0 10- 8 10- 6 10- 4 10- 2 1000
5
10
15
20
SerFBP
RLU
Serine 1
0mM
Lysine 1
0mM
Tryptophan
10mM
Tyrosin
e 10m
M
Threonine 1
0mM
Aspara
gine 10m
M
Glutamate
10mM
Basal
activ
ity
Alanine 1
0mM
Arginine 1
0mM
Histidine 1
0mM
Isoleu
cine 1
0mM
Leucin
e 10m
M
Aspart
ate 10
mM
Glutamine 1
0mM
noPEP
Glycine 1
0mM
Prolin
e 10m
M
Valine 1
0mM
Phenyla
lanine 1
0mM
Methionine 1
0mM
Cystin
e 10m
MFBP 50
uMFBP 50
uM
0
200
400
600PKM2
Of all proteinogenic amino acids only serine activates PKM2
Two known modifier amino acids of PKM2 (Tyr105 and Cys358) are in the vicinity of the serine binding domain
Acknowledgements
Marc O’ReillyPetra HillmannAgnès MartinJoe Coyle
Finn HoldingAdam Hold
Barbara ChanetonLiang (Leon) ZhengChristian FrezzaElaine MacKenzie
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