„Preparation of the concerned sectors for educational and R&D activities related to the Hungarian ELI project ”

THz spectroscopy in biology

Andrea Buzády

6. lectureSpectroscopy

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6. lecture

Spectroscopy – Biological applicationClassical / quantum chemical modelling

• Quantum chemicalmodelling: DensityFunction Theory (DFT)

• Classic chemicalmodelling: Chemistry atHARvardMacromolecularMechanics (CHARMM)

Vibrational and rotationalspectra of (simple) molecules can be derivedfrom different theoreticaland empirical modelling.

glicin

glicin

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6. lecture

Spectroscopy – Biological applicationClassical / quantum chemical modelling

Absorption peaks and related molecularmotions of simpleamino acid: glicin.

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6. lecture

Spectroscopy – Biological application

Terahertz spectroscopy of enantiopure and racemic polycrystalline valineMichael R. C. Williams, Alan B. True, Artur F. Izmaylov, Timothy A. French, Konstanze Schroecka and Charles A. SchmuttenmaerPhys. Chem. Chem. Phys., 2011,13, 11719-11730

Amino acids → peptides, proteins

D-valin

L-valin

DL-valin

T= 78KT=300K• Widening of peaks at room temperature

compared to low temperature (78 K)• L and D stereisomers have similar

spectra• DL racemat mixture has different

spectrum from D and L-valin• DL-valine does not split into multiple

peaks at low temperature, although it does sharpen

1.70 THz

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6. lecture

Spectroscopy – Biological application

Terahertz spectroscopy of enantiopure and racemic polycrystalline valineMichael R. C. Williams, Alan B. True, Artur F. Izmaylov, Timothy A. French, Konstanze Schroecka and Charles A. SchmuttenmaerPhys. Chem. Chem. Phys., 2011,13, 11719-11730

Amino acids → peptides, proteins

Temperature dependence of L- (and D-) valine THz spectra.Varied temperature shifts the absorptionpeaks related to different vibrationmodes in different manner.

The absorption coefficients (in cm-1) have been corrected and offset by the temperature at which the spectrum was collected (from 78 K to 298 K in steps of 20 K) for better presentation.

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6. lecture

Spectroscopy – Biological application

Terahertz spectroscopy of enantiopure and racemic polycrystalline valineMichael R. C. Williams, Alan B. True, Artur F. Izmaylov, Timothy A. French, Konstanze Schroecka and Charles A. SchmuttenmaerPhys. Chem. Chem. Phys., 2011,13, 11719-11730

Amino acids → peptides, proteins

Peak points (obtained by fitting Voigt functions to each spectrum on previous slide)Overlapping features between 1.5 and 1.9 THz consisted of three peaksfeature at 2.25 THz corresponds to a single peak

The 1.7 THz absorption peak does notchange, the others blue-shift withincreasing temperature

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6. lecture

Spectroscopy – Biological application

R. Balu, S. K. Gregurick, E. J. Heilweil, Determination of enantiomeric compositionby terahertz spectroscopy: mixtures of D-and L-tryptophan.

Amino acids → peptides, proteins

Spectra are shifted along Y axis for better representation!

THz absorption spectra for the pure D-and L-tryptophan (Trp) enantiomers, their 1:1 mixture (DL-tryptophan) and F6-fluorine-substituted DL-tryptophanamino acid in the solid state.• L-Trp and D-Trp stereoisomers

show identical spectra, whereas the DL-Trp mixture has a distinctly unique spectrum.

• The F6-DL-Trp also shows characteristic spectral features, different from the DL-Trp mixture.

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6. lecture

Spectroscopy – Biological applicationAmino acids → peptides, proteins

Spectra are shifted along Y axis for better representation!

THz absorption spectra ofseveral di- and tripeptides

variability and uniqueness for each structureLonger peptide (top spectrum of a random 10-mer) showns loss of distinguishable absorption peaks,

most likely from spectral overlap or hydrated water.

alanin valinFrequency (THz)

Abs

orba

nce

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6. lecture

Spectroscopy – Biological application

Applications of Terahertz Spectroscopy in BiosystemsDavid F. Plusquellic, Karen Siegrist, Edwin J. Heilweil, and Okan EsenturkChemPhysChem, 2007, 8, 2412 – 2431

Amino acids → peptides, proteins

Three different crystalline forms of trialanine.Although the conformational forms of the monomers are similar, the parallel and hydrated antiparallel β-sheets have different space groups (P12 and C2, respectively, the dehydrated form is unknown but likely (C2). These spectra indicate the sensitivity to the different space groups as well as the impact of the hydrogen bonded (structural) water.

2 structure of trialanine was known (X-ray crystallography). Third form discoveredin this study was the dehydrated form: ap-Ala3. Frequency (THz)

Abs

orba

nce

trialanines

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6. lecture

Spectroscopy – Biological application

Chemical Physics Letters 418 (2006) 65–70Terahertz spectroscopy of solid serine and cysteineT.M. Korter, R. Balu, M.B. Campbell, M.C. Beard, S.K. Gregurick, E.J. Heilweil

Amino acids → peptides, proteinsA single atom difference can resultin different THz absorbance• More definite absorption peaks at

low temperature (77 K)• An oxigen (O) → sulfur (S)

substitution of serine → cysteineresults in characteristic spectrum

• Most relevant difference in therange of 0-100 THz

1 single atom difference!

Spectra are shifted along Y axisfor better representation!

next

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6. lecture

Spectroscopy – Biological applicationAmino acids → peptides, proteins

Chemical Physics Letters 418 (2006) 65–70Terahertz spectroscopy of solid serine and cysteineT.M. Korter, R. Balu, M.B. Campbell, M.C. Beard, S.K. Gregurick, E.J. Heilweil

Spectra are shifted along Y axisfor better representation!

Low frequency range (0-100 THz) part of the absorbancespectra of L-serine and L-cysteineMost relevant difference is at 66 THz (peak of serine)

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6. lecture

Spectroscopy – Biological application

Applications of Terahertz Spectroscopy in BiosystemsDavid F. Plusquellic, Karen Siegrist, Edwin J. Heilweil, and Okan EsenturkChemPhysChem, 2007, 8, 2412 – 2431

Amino acids → peptides, proteins

3D structure of myoglobinFrequency (THz)

Abs

orba

nce

High alfa helical(secondary struc-ture) content of myoglobin doesnot result in helixspecific absorban-ce spectra.

Multitude of amino acids in the chain results in overlapping absorption peaks, that leads to non-characteristic spectra. No peaks can be found.

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6. lecture

Spectroscopy – Biological applicationAmino acids → peptides, proteins

A. Markelz, S. Whitmire, J. Hillebrecht, R. Birge, Phys. Med. Biol. 2002, 47, 3797.

• Longer amino acid chain → more vibration• More collective vibration → more crowding and

overlapping in the spectrum• Result: non-characteristic, featureless spectra

linearly increasing absorbance at low THz range(0-2 THz)

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6. lecture

Spectroscopy – Biological applicationNucleic acids → DNA, RNA

Nucleic acids consist of one nucleobase from 5 possible (A,G,T,C,U)Nucleobase binds either to ribose or deoxyribose, in case of RNA or DNA, respectively → (deoxy)ribonucleoside is formedFurther binding of 3 phosphates results in nucleic acid, like ATP, GTP, …

Is there any simpleway to devine thespectra of a nucleicacid from thespectra of puresacharose (ribose) and nucleobaseand phosphate? NO!

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6. lecture

Spectroscopy – Biological applicationNucleic acids → DNA, RNA

Double stranded DNA absorption depends onhumidity of air.The coiled-coil structureof DNA varies with thehumidity as hydrogenbonding involving watermolecules are more orless frequent underdifferent conditions.

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6. lecture

Spectroscopy – Biological application(poly)saccharides

1

1

2

2

Protonated (H) and deuterated (D) sucrose spectra at differenttemperature shows varied positionof absorbance peaks.

Blueshift at low temperatures, then redshift after a critical temperature.

next

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6. lecture

Spectroscopy – Biological application(poly)saccharides

Temperature induced shift of peaksRelative shift:

protonated and deuteratedpolycrystalline sucrose(from 10 to 300 K)

f(T)/f(10K)-1

Blueshift at low temperatures, then redshift after a critical temperature.

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6. lecture

Spectroscopy – Biological application(poly)saccharides

M. Walther, B. M. Fischer, P. Uhd Jepsen, Chem. Phys. 2003, 288, 261.

THz absorption spectra of glucose and fructose.

Two simple sugar incrystal form.

Typical widening of spectral peaks athigh temperature.

Characteristicpeaks related to thedifferent sugarmolecules.

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6. lecture

Spectroscopy – Biological application(poly)saccharides

M. Walther, B. M. Fischer, P. Uhd Jepsen, Chem. Phys. 2003, 288, 261.

Simple sugar of sucrose in crystalform and glucose(now in amorphouspowder form).

Unstructuredspectra of non-crystal (amorph) form sugars.

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6. lecture

Spectroscopy – Biological application(poly)saccharides

P.C. Upadhya, Y. C. Shen, A. G. Davies, E. H. Linfield, Vib. Spectrosc. 2004, 35, 139.

The THz spectra of (a) D-glucose, (b) D-mannose, (c) D-galactose and (d) D-fructose• Widening of peaks at high temperature

compared to peaks at low temperature (4 K)• At high temperature some specific peaks

present at low temperature disappear.

Measured at 4 and 295 K. The spectra at 4 K are vertically offset for clarity.

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6. lecture

Spectroscopy – Biological application(poly)saccharides

P.C. Upadhya, Y. C. Shen, A. G. Davies, E. H. Linfield, Vib. Spectrosc. 2004, 35, 139.

THz absorption spectra of (a) maltose and (b) lactose.

The spectra at 4 K are vertically offset for clarity.