1. Characterizing Peptide β-Hairpin Loops via Cold Ion Spectroscopy of Model Compounds
John T. Lawler, Andrew DeBlase, Chris Harrilal, Scott A. McLuckey, Timothy S. Zwier

2. Pancera, M.; et.al. Nature 2014, 514 (7523), 455–461.
Why study β-turns?
Ubiquitous in biology and often used in drug/synthetic design
Turns provide 3D structure via allowing the protein to fold over itself
Diener, M. E., et.al., . J. Am. Chem. Soc. 2015, 137 (38), 12369–12377.
Pancera, M.; et.al. Nature 2014, 514 (7523), 455–461.
Marcelino, A. M. C. , et.al. ,Biopolymers 2008, 89 (5), 380–391.

3. Steric Effects and Differences in Structure
YAPAA DP LP
syn
anti
C11,C17,πNH/C14/trans/FNH/C10/C7OH
C5,C11,πNH/C14/trans/FNH/C10/C7OH
3.15 kJ∙mol-1
35.70 kJ∙mol-1
What happens when the residue
C-terminal to proline is less bulky?
YGPAA

4. Size and β-turn formation
Will increasing the size of the peptide lock in the β-turn ?
Will we see the beginnings of beta sheet formation?

5. Instrumentation Spectroscopy axis MS axis q3 q2 q1 Nano ESI Det 1
qtrans2
qtrans1
Cold
Trap
Turning Quad 1
Turning Quad 2
Nano ESI

6. Question 1
Does removing steric hindrance C-terminal to Proline change the proclivity for β-turns?

7. UV Photofragmentation Spectroscopy
YADPGA
Reduced number of conformers!
YALPGA
Changed this from side by side spectra, not really liking it

8. YADPGA IR-UV Holeburning Spectrum
Holeburn Spectrum
*IR fixed on cm-1
YADPGA
IR-UV holeburning can be used to obtain conformation-specific UV spectra of ions. We fix the IR laser on an infrared transition in the IR spectrum and scan the UV probe. Any UV transitions sharing the same ground state as the IR transition will show a depletion in photofragment signal.
Wavenumber (cm-1)
One major conformer is seen, minor conformer(s) are likely present

9. Glycine Position Effecting Conformational Preferences
Gly-substituted D-Proline peptides show a reduced amount of conformers
(a) YADPGA
YGLPAA
(a) YALPGA
Gly-substituted L-Proline peptides present with an increased number of conformers

10. IR Gain Spectroscopy of YADPGA
C10 Hydrogen Bond
*Calculations performed at the B3LYP-GD3BJ/6-31+G* level of theory
C10
NH3+ a
in addition to the depopulation of the ground state, absorption of a resonant IR photon can lead to intramolecular vibrational energy redistribution (IVR). This redistribution of energy among accessible vibrational levels results in statistical inhomogeneous broadening of the UV spectrum to the red of the electronic origin.  Thus, in IR-UV gain spectroscopy, the UV laser is fixed at a wavelength to the red of the lowest energy electronic origin in the UV spectrum. Vibrational excitation acts to induce UV absorption at this wavelength, resulting in a gain in UV photofragmentation signal.
C14
Tyr OH
Free NH
C14 Hydrogen Bond
C10 and C14 H-bonds are diagnostic for β-turn

11. Question 2
Will increasing the size of the peptide change the proclivity for β-turns?

12. Enlarging the peptide reduced the number of conformers to one
YAADPGAAA UV Spectrum
YAADPGAAA
YADPGA
Enlarging the peptide reduced the number of conformers to one

13. YAADPGAAA Infrared Spectra
Amide II
Amide I
Hydride Stretch
-NH3+ a
C101/C102/C7
-NH3+ a
C14
-NH3+ a
Free NH
Acid OH
Tyr OH
CH Stretches

14. Calculated Lowest Energy/Best Fit Structures
C10 Hydrogen Bond
C14 Hydrogen Bond
*Calculations performed at the B3LYP-GD3BJ/6-31+G* level of theory

15. Conclusions
Does removing steric hindrance C-terminal to Proline change the proclivity for β-turns?
YES, UV action spectroscopy and IR-UV holeburning confirmed a reduced number of conformers when GLY was placed C-terminal to D-Proline
In the case of L-Proline the inverse was witnessed, more conformers are present
Will increasing the size of the peptide change the proclivity for β-turns?
YES, the peptide YAADPGAAA revealed a single conformer that forms the expected beta-turn
Will we see the beginnings of beta sheet formation?
NO, increasing the size of the peptide did not produce the tell-tale signs of a beta sheet

16. Future Work
Complete Infrared depletion spectroscopy experiments on YAADPGAAA , YADPGA, and YALPGA
Do a more exhaustive search of possible structures for each of the experimental peptides

17. Acknowledgements Jonathan Amy Facility for Chemical Instrumentation
National Science Foundation
Zwier Group
McLuckey Group