Picosecond primary structural transition of the heme is retarded after nitric oxide binding to heme proteins

Abstract

We investigated the ultrafast structural transitions of the heme induced by nitric oxide (NO) binding for several heme proteins by subpicosecond time-resolved resonance Raman and femtosecond transient absorption spectroscopy. We probed the heme iron motion by the evolution of the iron-histidine Raman band intensity after NO photolysis. Unexpectedly, we found that the heme response and iron motion do not follow the kinetics of NO rebinding. Whereas NO dissociation induces quasi-instantaneous iron motion and heme doming (<0.6 ps), the reverse process results in a much slower picosecond movement of the iron toward the planar heme configuration after NO binding. The time constant for this primary domed-to-planar heme transition varies among proteins (∼30 ps for myoglobin and its H64V mutant, ∼15 ps for hemoglobin, ∼7 ps for dehaloperoxidase, and ∼6 ps for cytochrome c) and depends upon constraints exerted by the protein structure on the heme cofactor. This observed phenomenon constitutes the primary structural transition in heme proteins induced by NO binding.time-resolved Raman spectroscopy | allostery | structural dynamics T he role of diatomics endogenously generated in cells as messengers in various signaling pathways has been demonstrated for nitric oxide (NO) (1, 2) and carbon monoxide (CO) (3, 4) and various gas-sensors heme proteins have been discovered in the last two decades (5). In these signaling pathways, the binding of diatomics to their respective heme sensor domain triggers a response of the associated enzymatic domain by modulating the catalytic or the gene regulating activity (5). Recently, myoglobin (Mb) was proposed to be involved in the regulation of NO level (6), and neuroglobin was proposed as a sensor regulating the NO∕O 2 balance (7). Similarly to Hb (8-10), the signaling mechanisms are based at the molecular level on an allosteric structural change of the receptor/sensor protein coupled to a change of activity or affinity and are induced by diatomic ligation to the heme iron cofactor. The signal of diatomic binding/release can be transmitted to the overall protein structure either by in-plane/out-of-plane motion of the iron through the bound proximal histidine or by the cleavage of the iron-histidine (Fe-His) bond. In particular, for Hb and its monomeric counterpart Mb, the out-of-plane iron motion occurs in less than 1 ps after gaseous ligand release (11,12) and represents the first event of the allosteric R → T transition (9). This primary heme iron motion is followed by overall tertiary changes, notably motion of α-helices (13). Upon gaseous ligand binding, the reverse T → R allosteric transition is coupled to the iron motion toward the heme plane, and this primary motion is also considered quasi-instantaneous since the stereochemical description of Hb allostery by Perutz (9, 10) although up to now it has never been observed.The ultrafast events in the interaction of heme proteins with various gaseous ligands were studied extensively at molecular level in the last two decades by femtosec...

Motion of proximal histidine and structural allosteric transition in soluble guanylate cyclase

“…Because the Fe-His has been reformed, we assign this decay to NO rebinding to the 5c-His. However, this time constant is much larger than those time constants measured for geminate rebinding of NO to 5c-His from within the heme pocket of myoglobin, hemoglobin, and dehaloperoxidase (31). In sGC, NO is expected to have already escaped the heme pocket after several nanoseconds but not to have diffused to the solvent, so that τ 3 must reflect rebinding from the protein core.…”

“…After the 5c-His heme is reformed, the subsequent NO rebinding occurs two orders of magnitude more slowly (τ 1 = 6.5 ns) without any component in the 100-to 200-ps time range, as found for the globin family (31), or in the 200-to 400-ps time range for endothelial NO-synthase (38). However, nanosecond NO rebinding components were already observed in the case of NO-synthase (38) and have been assigned to NO already present within the protein access pathway from solvent.…”

Reproducing kernel potential energy surfaces in biomolecular simulations: Nitric oxide binding to myoglobin

“…This will be of particular interest in better characterizing the rebinding dynamics of photolyzed NO for which recent experimental work suggested the existence of a NO-bound-like, Fe-out-of-plane metastable state. 7 Reactive molecular dynamics simulations 76,77 involving the present 2 A and 4 A kernel-based PESs will provide the necessary information for an atomistically resolved picture linking experiment and molecular dynamics.…”

“…7 This work points towards a direct coupling between these degrees of freedom on the 10 to 100 ps time scale. Running a statistically significant number of QM/MM trajectories from which to analyze and atomistically resolve the interplay of the motions involved is beyond current computational methods.…”

“…1 The rebinding kinetics of NO to Mb has been studied intensively over the past three decades using experimental and computational methods. [2][3][4][5][6][7][8][9][10][11][12][13][14] Compared to rebinding of CO, no substantial rebinding barrier has been found for geminate recombination of NO to the heme Fe 3 and ab initio calculations have even suggested that the recombination reaction may be barrierless for NO in specific conformations. 15 Because of the short time scale involved (picoseconds), this process is ideally suited to be investigated computationally at an atomistic level.…”