Ultrafast dynamics in photoexcited Neuroglobin

revealedby Femtosecond Stimulated

Raman Scattering

Autore:Giovanni Batignani

Relatore:Tullio Scopigno

Emoproteine e Neuroglobina

Il nucleo di ogni emoproteina è costituito da uno ione di ferro,

che ne determina le funzionalità biologiche.

Heme

Ligando gassoso (O2, CO, NO)

Ferro

Amminoacidi

Funzione biologica ↔ formare e rompere legami

Specie deoxy pentacoordinata

Specie deoxy esacoordinata

Dinamica Ultraveloce

Tecnica Pump ProbeStudiare dinamiche di campioni fotoeccitati

∆t<1ps

Dinamica Ultraveloce

Time Resolved Resonance Raman (TR3)Citocromo cNeuroglobina deoxy

????

Fotolisi della Metionina

Stati transienti ??

δω≈10cm-1 δt=2ps

Jean-Louis Martin et al. 2007“…However, such studies are extremely challenging for the physiologically relevant ligand oxygen and have not been reported yet, even for the well studied oxygen carriers myoglobin and hemoglobin, presumably at least in part because of technical difficulties”

“…Because of the decreased spectral resolution in the subpicosecond Raman experiments, lines in these spectra are broader than in the steady-state spectra obtained with cw excitation.”

Dinamica Ultraveloce

Assorbimento Transiente (TA)

“Transient spectra indicate the formation of a 5-c domed species upon photoexcitation of the 6-c species, and thus photodissociation of an internal residue. The process appears similar to the way small ligands like CO can be photodissociated from ferrous heme-His-CO complexes. As, in contrast to small gaseous ligands, the internal ligands are restrained to heme proximity, full rebinding is observed on the picosecond time scale for all proteins.”

Femtosecond Stimulated Raman Scattering

Raman Pulse

Ram

an P

ulse

Raman Pulse

Timeline

Raman ProbeR

aman Probe

Raman spontaneo“transform limited”

δωδt>15ps·cm-1

…+𝒆

− 𝒊(𝝎

𝑹−𝝎

𝑷)𝒕

Phot

oche

mic

al P

ump

Ener

gy

“Reaction” coordinate

Actinic Pum

∆t

Risoluzione temporale illimitata

Timing cycle – 1 ms timebase:

WLC ON ON ON ON ….

RAMAN PULSE ON OFF ON OFF ….

PHOTOCHEM PUMP ON ON OFF OFF ….

Raman Pulse

Ram

an P

ump

WLG WL Probe

FSRS SetupFemtosecond Stimulated Raman Scattering

Femtosecond Stimulated Raman Scattering

Conclusioni

Risultati ottenuti

Conclusioni

Prospettive di ricerca future