Transcript YangzhongQin Spectroscopy Symposium OSU 06212013 submit.pptx

Using Tryptophan As A Probe For Studying the Protein
Hydration Dynamics
Yangzhong Qin
June 21st 2013
The Ohio State University
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Background:
• Tryptophan has been using as an optical
probe to study solvation dynamics for
decades, especially for protein dynamics.
•Tryptophan has large dipole moment
change from excited state to ground state.
•The detection of the initial ultrafast
dynamics is limited by the instrument
response.
?
349nm
•Tryptophan in water has two lifetimes,
which are 0.5ns and 3.0ns.
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Methodology: Fluorecence Upconversion
 Fix the BBO and detector, move the delay stage -------- 2D FRES
 Fix the delay, move the BBO and dectector -------- 3D FRES
I  (t )  I solv (t )  I popul (t )   ai et /  i   b j e
i
 (t )   ()
C (t ) 
 (0)   ()
t /  j
j
C (t ) 
 s (t )   l (t )
 s ( 0)   l ( 0)
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Coumarin mixture mimic tryptophan
Coumarin mix 5μM
C152/C153 (50:50)
in methanol.
pump@400nm
λSS = 534nm
λRaman = 455nm
ν0 =21710*
ν0 =21266, λ0 = 463.8nm
ν0 =20835, λ0 = 475.3nm
2ps (844cm-1)
13ps (1252cm-1)
The bluer, the better!
* Bose et al., J. Phys. Chem. B 2009, 113, 11061–11068
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Simulation of initial fast component loss
 p (t)  (    )0.5 * e-t/  0.5 * e-t/  

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 For fast dynamics, after Instrument
response time (FWHM), almost no missing
of the initial fast component.
 For slow dynamics, missing part is
negligible.
 For an example , if the instrument response
time FWHM~0.4ps,and the fast dynamics is
0.3ps, the instrument can recover 75% of
such fast dynamics. However, this is
without deconvolution, which means it sets
the missing percentage uppper limit.
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Tryptophan Initial fast component loss
Pump @
ν0 (direct)
ν0 (deconvoluted)
ν0 (true)
Missing
280nm
31046cm-1
(318.8nm)
31515cm-1
(315.4nm)
31840cm-1
(312.2nm)
325cm-1
290nm
30809cm-1
(321.4nm)
31141cm-1
(319.1nm)
31210cm-1
(318.2nm)
69cm-1
295nm
30511cm-1
(324.5nm)
30754cm-1
(322.6nm)
30895cm-1
(321.3nm)
141cm-1
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“True ” value are extrapolated from data at 2K (Scott et al., J, M. Chem. Phys. 1989,131, 63-79)
Lifetime contribution to the solvation correlation function
C (t ) 
 (t )   ()
 (0)   ()
C (t ) 
 s (t )   l (t )
 s ( 0)   l ( 0)
Multiple lifetime components will make
contribution to the dynamic stokes
change. It is necessary to consider such
contribution, especially for slow dynamics.
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Slowdown of the protein solvation dynamics (1)
N35W
F30W
A1
0.320
E1 1296
A1
0
E1
0
A2
0.169
E2
685
A2
0.291
E2
177
A3
0.038
E3
155
A3
0.205
E3
125
A4
0.047
E4
189
A4
0.503
E4
306
A5
0.426
E5 1724
A5
1.285
E5
781
V0
30630
E
V0
31654
E
1389
2326
Vinf 26579
Vinf 30264
T1
0.340
T1
NA
T2
2.050
T2
2.40
T3
16.770
T3
13.5
T4
154.2
T4
171.6
T5
1624
T5
1160
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Slowdown of the protein solvation dynamics (2)
C (t ) 
 s (t )   l (t )
 s ( 0)   l ( 0)
Summary:
Tryptophan can be a powerful probe for detecting
protein hydration dynamics, but time deconvolution
and lifetime contribution need to be considered.
Solvation water around protein has been
significantly slowdown due to water-protein
interaction.
Sample
T1
T2
T3
T4
A1%
A2%
A3%
A4%
Total E (cm-1 )
Trp in water
0.34
1.5
NA
NA
49.6
50.4
0
0
2407
N35W
0.34
1.8
15.8
222.5
52
30.5
8
9.5
2397
F30W
NA
2.4
16.2
275.3
0
21.3
15.1
63.6
871
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Thank you!
Dr. Chihwei Chang
Dr. Jeffray Stevens
MS. Lijuan Wang
Advisor: Dr. Dongping Zhong
Funding from:
NSF
NIH
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