2 research outputs found
Electronic and Nuclear Structural Snapshots in Ligand Dissociation and Recombination Processes of Iron Porphyrin in Solution: A Combined Optical/X-ray Approach
The
photodissociation and recombination of CO and 1-methylimidazole
(Im) from iron protoporphyrin IX (FePP–ImCO) dissolved in a
30% v/v aqueous solution of Im was studied using ultrafast optical
transient absorption (TA) and X-ray transient absorption (XTA) spectroscopies.
FePP–ImCO was shown to lose the CO ligand upon excitation at
the Q bands, with 3.8 ps vibrational cooling and 21.6 ps intersystem
crossing time constants derived from optical TA experiments, followed
by ligation of a second Im on the nanosecond time scale. The penta-coordinate
FePP–Im intermediate which forms following CO dissociation
adopts a square pyramidal geometry with a “domed” iron
center that is reminiscent of that formed upon loss of CO from carbonÂmonoxyÂmyoglobin
(MbCO). Unlike MbCO, which typically retains its newly generated penta-coordinated
geometry until CO recombination, FePP can adopt a hexa-coordinate
geometry by binding an additional Im ligand (FePP–(Im)<sub>2</sub>), allowing the porphyrin to exist in the low-spin electronic
state even without the CO attached. The second Im ligand remains bound
until CO recombination occurs with a time constant of 283 ÎĽs.
The photodissociated states of FePP–ImCO and MbCO 100 ps after
photoexcitation have similar iron site geometries, implying that the
protein matrix in MbCO maintains minimum potential energy in the heme
center despite the large-scale reorganization in the protein secondary
and tertiary structure that arises from the dynamic active site/matrix
interaction
Megahertz pulse trains enable multi-hit serial femtosecond crystallography experiments at X-ray free electron lasers
The European X-ray Free Electron Laser (XFEL) and Linac Coherent Light Source (LCLS) II are extremely intense sources of X-rays capable of generating Serial Femtosecond Crystallography (SFX) data at megahertz (MHz) repetition rates. Previous work has shown that it is possible to use consecutive X-ray pulses to collect diffraction patterns from individual crystals. Here, we exploit the MHz pulse structure of the European XFEL to obtain two complete datasets from the same lysozyme crystal, first hit and the second hit, before it exits the beam. The two datasets, separated by <1 µs, yield up to 2.1 Å resolution structures. Comparisons between the two structures reveal no indications of radiation damage or significant changes within the active site, consistent with the calculated dose estimates. This demonstrates MHz SFX can be used as a tool for tracking sub-microsecond structural changes in individual single crystals, a technique we refer to as multi-hit SFX