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Towards the Concise Syntheses of Selenium- and Tellurium-Containing Tryptophan Analogs for the Elucidation of Protein
Structure and Function Ryan Agh§¶, Ambrose Rice§¶, Duane M. Hatch§¶, Ricardo Martí-Arbona¶, L.A. “Pete” Silks¶
¶Bioscience Division, Group B11, Los Alamos National Laboratory, Los Alamos, NM, 87545 USA §Department of Chemistry & Physics, Belmont University, Nashville, TN, 37212
Bioscience
Innovation in
Health and Security
1. Welch, M.; Philips, R.S., Heterocyl. Com., 1999, 5, 305-310.
2. Hatch, D. M. et al. Current Org. Chem. 2004. 47-64.
3. Hatch, Duane. Tenn. Tech. Univ. 2003. pp. 4-8, 19-70.
4. Boles, J.O.; Henderson, J.; Hatch, D.; Silks, L.A. Biochem.
Biophys. Res. Comm. 2002. 257-261
References
Conclusions
• We have successfully validated our proposed synthesis
• We have analyzed all intermediates and products via 1H, 13C, 77Se, and 125Te NMR spectroscopy.
• We have successfully bio-incorporated SeTrp into a model
protein.4
Acknowledgements • Scott Robbins (LANL)
• Kirill Balatsky (LANL)
• Katrina Barnett (Belmont University)
• Belmont University Department of Chemistry and Physics
• Bioscience Division, B-11, LANL
• Partial funding from DOE Visiting Faculty Program 2015
• LANL Student Programs Office
Benefits
• Increase the number of available unnatural amino acids for
protein structure/function elucidation.
• Tryptophan is often found near or leading into the active
site of a protein, thereby allowing for interrogation of the
environment around that residue by NMR spectroscopy.
• Se-Trp and Te-Trp are less fluxional than Se-Met giving
rise to increased stability.
Future Work
• Continue scaling up the synthesis
and refining methods.
• Characterize catalytic activity of
Lignin Peroxidase with Trp
analogs.
• Improve stability of intermediate
compounds
• Get crystallographic data and
resolve the 3D structure of
selected proteins.
• Investigate catalytic activities of
SeTrp and TeTrp containing
proteins.
Unclassified 'LA-UR-15-25780'
Background
Since the 1990’s, Se-Methionine (Se-Met) has been invaluable in
the elucidation of 3D-protein structures. There have also been
published results of a successful synthesis of Se-Trp (Scheme
1).1 However, this synthetic route is an expensive one, with low
Scheme 1: Phillip’s Modified
Paulmier Synthesis1
yield, and the
inability to label the
compound with the 77Se isotope. Our
synthetic strategy is
more refined and
allows for 77Se
insertion. (Scheme
2).2,3
1. Determine the viability of using
this route for large production of
6H-selenolo[2,3-b]pyrrole.
2. Synthesize 6H-telluro[2,3-
b]pyrrole utilizing this method.
3. Maximize the enzymatic coupling
of the serine side chain using
purified Tryptophan Synthase.
X = Se or Te
Scheme 2: LANL Synthesis2,3
Goals
Trp
Synthase
Figure 1: L-Tryptophan
Figure 2: [6,7]SeTrp or [6,7]TeTrp
Overview
We worked on an improved synthesis which forms an amino
acid (L-Tryptophan, Figure 1) analog. Our synthesis can be
used to insert either Selenium or Tellurium into the amino acid
(Figure 2). This was done to investigate protein structure and
catalytic activity via X-Ray Crystallography (Figure 3) and
NMR Spectroscopy.
89.00%
10.00% 0.70%
0.30%
X-Ray
Crystallography
NMR Spectroscopy
Electron
Microscopy
Other Methods
Figure 3: Methods for Solving Protein Structure X = Se or Te
No Hh, Hi, or Hj
Syntheses
6H-selenolo[2,3-b]pyrrole 6H-telluro[2,3-b]pyrrole
63% Yield
82% Yield 75%Yield
80% Yield
80% Yield
He
Hg
Hd Hc
Hb
Ha
2.2 2.4 2.6 2.8 3.0 3.2 3.4 3.6 3.8 4.0 4.2 4.4 4.6 4.8 5.0 5.2 5.4 5.6 5.8 6.0 6.2 6.4 6.6 6.8 7.0 7.2 7.4 7.6 7.8 8.0
f1 (ppm)
1 Hj
Hh
Hi
2.2 2.4 2.6 2.8 3.0 3.2 3.4 3.6 3.8 4.0 4.2 4.4 4.6 4.8 5.0 5.2 5.4 5.6 5.8 6.0 6.2 6.4 6.6 6.8 7.0 7.2 7.4 7.6 7.8 8.0
f1 (ppm)
1
Hj
Hh
Hi
Hf
He
Hg
Hf
77Se-NMR Chemical Shifts
No He, Hf, or Hg
169.97 ppm 507.83 ppm 437.44 ppm
No Hh, Hi, or Hj