Euromar2009 talk
17
CORRELATION SPECTROSCOPY IN ASCORBATE OXIDASE USING HIGH- FIELD PULSED EPR Alexey Potapov Weizmann Institute of Science
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CORRELATION SPECTROSCOPY IN ASCORBATE OXIDASE USING HIGH-FIELD PULSED EPR
Transcript of Euromar2009 talk
CORRELATION SPECTROSCOPY IN ASCORBATE OXIDASE USING
HIGH-FIELD PULSED EPR
Alexey PotapovWeizmann Institute of Science
MW
RF
echo
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a
ff
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MW RF
ENDOR- electron-nuclear double resonance
Double resonance experiments.
MW2
MWFID
MW
RF
echo
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a
ff
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|N> |N>
MW RF
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a
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MW2
ELDOR- (electron-electron double resonance) – detected NMR
ENDOR- electron-nuclear double resonance
ELDOR-detected NMR and ENDOR.
ELDOR-detected NMR
ENDOR
IntensityDepends on forbidden electron
transitions
Depends on allowed nuclear
transitions
ResolutionLow (determined by MW field strength,
but Tm limited)
High (determined by the strength of
RF field B2 )
NucleiPreferable for broad lines 14N,55Mn,61Ni
Narrow lines
ELDOR-detected NMR and ENDOR.
Good resolution at high-field
TRIPLE ENDOR
MW
RF1
RF2
echo
MW2
RF
MWFID
THYCOS
THYCOS: correlating ELDOR-detected NMR and ENDOR spectra
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f
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|12>a
MW
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a(n)|12>
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RF
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f
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MW
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a(n)|12>
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RF
MW2
RF
MWFID
Ms |>
Ms |>
e
n1
n2
A1
A2
THYCOS: correlating ELDOR-detected NMR and ENDOR spectra
variable
-> nuclei belonging to the same paramagnetic center >-the lines from the opposite electronic manifold
12
A1 A2
ED-NMR excitation
I12
A1>0 A2>0
I12
A1>0 A2>0
THYCOS: correlating ELDOR-detected NMR and ENDOR spectra
e
n1
n2
A1
A2
N
N2
H
Him
H
H
COOH
Ha
N1
Ham
Ham
H
Cu
N
N2
H
Him
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H
COOH
Ha
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Ham
Ham
H
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H
H
HH
-10 -8 -6 -4 -2 0 2 4 6 8 10-10 -8 -6 -4 -2 0 2 4 6 8 10
H-
RF, MHz
ENDOR
THYCOS
Frozen solution: complex of Cu2+ L-histidine
-120-100 -80 -60 -40 -20 0 20 40 60 80 100 120
Frequency, MHz
N1,2
HTA
dq N1,2
3.00 3.05 3.10 3.15 3.20 3.25 3.30 3.35
Magnetic field, T
Type 1
-protons
Frozen solution: ascorbate oxidase
Type 2
S
3000 3100 3200 3300 3400
Magnetic field, mT
B0=3360 mT
B0=3300 mT
B0=3200 mT
B0=3075 mT
3000 3100 3200 3300 3400
Magnetic field, mT
B0=3360 mT
B0=3300 mT
B0=3200 mT
B0=3075 mT
B0=3360 mT
B0=3300 mT
B0=3200 mT
B0=3075 mT
Frozen solution: ascorbate oxydase
-20 -15 -10 -5 0 5 10 15 20
RF-v1H, MHz
Axx,Ayy,Azz=[30 30 36] MHz and Euler angles (()=(0, 50, 0)
3360
3330
3200
3075
B0,, mT
Frozen solution: ascorbate oxydase
-60 -40 -20 0 20 40 60
, MHz
14N1 sq
14N2, sq
14N1 dq
3360
3330
3200
3075
B0,, mT
Lines at ~10,~30 and ~20 MHz
Frozen solution: ascorbate oxydase
-60 -40 -20 0 20 40 60
, MHz
14N1 sq
14N2, sq
14N1 dq
3360
3330
3200
3075
B0,, mT
HYSCORE
t1 t2t1 t2
sqsq
dq
sqsq
dq
sqsq
dq
sqsq
dq
Lines at ~10,~30 and ~20 MHz
Frozen solution: ascorbate oxydase
-60 -40 -20 0 20 40 60
, MHz
14N1 sq
14N2, sq
14N1 dq
3360
3330
3200
3075
B0,, mT
=|A/2±I|I =20 MHz
Lines at ~10,~30 and ~20 MHz
sq-dq
sq-sq
sq-sqsq-dq
sq-sq
sq-sq
1, MHZ
2,
MH
Z
-60 -50 -40 -30 -20 -10 0 10 20 30 40 50 60
, MHz
A(-proton)>0 A(14N)>0 orA(-proton)>0 A(14N)>0
14N
ELDOR-detected NMR
Frozen solution: ascorbate oxydase
120 126 132 138 144 150 156 162
RF, MHz
x20
(A)
(B)
120 126 132 138 144 150 156 162
RF, MHz
x20
(A)
(B)THYCOS
ENDOR
Advantages:•Experiment is suitable for sampleswith broad lines and forbidden transitions.•Does not require large power.•No subtraction
Disadvantages:•Lack of resolution of ELDOR-dimension•Limited sensitivity.•Cavity bandwidth
THYCOS: correlating ELDOR-detected NMR and ENDOR spectra
Conclusions
•Using THYCOS experiment the lines are assigned as: Aiso=23 MHz – type 1; Aiso~40 MHz – type 2•Resolving species using spectroscopy only.
EPR team at Weizmann
Boris Epel - SpecMan
MW bridge – Yakov Lipkin, Yehoshua Gorodetski ,Koby Zibzner
Acknowledgements
