Rhodopsin, a pigment in the photoreceptor cells of the retina, absorbs light that enters the eye, transforming it into the first chemical signal in the chain reaction of vision.
Now researchers have zeroed in on the molecular dynamics that make this ultrafast reaction possible.
Scientists have long explored the chemistry that makes vision possible. But the primary chemical reaction underlying vision occurs so fast that details about what happens at the molecular level have remained elusive. This experiment investigated what particular movements the retinal molecule makes when light hits it, and how these molecular vibrations work in sync to initiate the first step in the chain reaction of vision.
The body uses a type of molecule called rhodopsin to detect light that enters the eye. The surfaces of the photoreceptor cells in the retina are dense with rhodopsin molecules embedded in the plasma membranes. Each rhodopsin molecule consists of the protein opsin and the chromophore retinal.
When light hits the retinal chromophore, it causes the molecule to isomerize, changing its shape and the shape of the entire rhodopsin molecule along with it. In this new state, rhodopsin can interact with other proteins in the photoreceptor cell, kick-starting the visual signaling cascade that ultimately sends a neural impulse to the brain.
During this key isomerization reaction, the retinal molecule flips like a switch from a cis- to a transorientation around its carbons 11 and 12, which are connected by a double bond .
This flip is incredibly fast. Because of its speed, previous research has been unable to identify the vibrations of the molecule that enable it. Chemical reactions like this one occur as a result of movements of the molecule that bring particular atoms into contact at just the right time, causing them to react to each other.
This experiment demonstrates that the primary chemical reaction of vision happens much faster than previously thought, and reveals the molecular dynamics that cause it.
These molecular movements can include twisting and stretching of the bonds between the molecule’s atoms, and many other types of movement. Each molecular movement emits a vibration that can be detected.
However, over the isomerization reaction of retinal, the molecule emits many different vibrations caused by many different movements of the molecule. Previous experiments didn’t examine the reaction on a short enough timescale to distinguish which of these vibrations were involved in the reaction and which were background noise.
This study used ultrafast heterodyne-detected transientgrating spectroscopy, exciting the retinal in rhodopsin with pulses of light to examine the isomerization reaction on a shorter timescale than ever before.
The researchers were able to characterize what happens to the retinal molecule during its isomerization at an unprecedented level of detail. By observing the event to a finer degree than previous studies, they captured a better picture of how long the reaction takes and pinpointed the particular vibrational modes that make it happen.
The researchers found evidence that the isomerization of retinal is a vibrationally coherent photochemical process with three particular vibrations that occur simultaneously in different parts of the molecule. These oscillate in sync to cause a single, ultrafast molecular flip.
They also found that the isomerization event happens much faster than previously thought, on a timescale of about 30 femtoseconds — that’s 30 millionths of a billionth of a second. Previous experiments had estimated the reaction to occur over about 200 femtoseconds.
The researchers concluded that the signals detected at 200 femtoseconds in previous studies had been due to the retinal becoming “vibrationally hot” during its recovery after isomerization, not to the isomerization reaction itself.
Three main reactive nuclear dynamics within the molecule appeared to drive the reaction. These movements were local stretching of the C11=C12 double bond, HOOP wags in and out of the local polyene chain plane by the C11 and C12 hydrogens and torsional vibration around the C11=C12 double bond.
The researchers used ultrafast heterodyne-detected transient-grating spectroscopy to observe the isomerization of retinal in a sample of bovine rhodopsin that had been isolated from frozen bovine retinae.
They used an anamorphic-pumped non-collinear optical parametric amplifier (NOPA) to generate pulses of light in the blue-green spectrum that lasted approximately 11 femtoseconds each.
Four ultrashort pulses of light were shot at the sample to excite its molecules. This caused the retinal to isomerize, and enabled the researchers to pick up the resulting resonances emitted by the retinal molecules.
The first two pulses of light were sent at the same time, striking the retinal sample simultaneously adjacent to one another and causing a standing wave interference pattern. The third pulse was shot to probe this interference pattern and a larger fourth pulse was used to detect the resulting interference.
Ultrashort pulses of blue-green light were used to observe the isomerization of the retinal molecule when it’s exposed to light — a key chemical process that drives vision — at a level of detail never before observed.
The experiment relied on the principle that as the retinal molecules became excited by the light pulses, their vibrations would alter an interference pattern these pulses created. Each type of vibration in the molecule that occurred would modulate the detected signal differently depending on its location, timing and type.
Because the transient-grating spectroscopy picked up many molecular vibrations at once, the signal was separated using Fourier filtering and time-domain analysis. The different types of vibrations were then identified and assigned to particular molecular movements at particular times.
The three major types of molecular vibrations were found to occur together at 30 femtoseconds in an unusually coherent manner.
This experiment has uncovered a detailed profile of the nuclear dynamics that happen in the primary chemical reaction of vision, providing a more complete explanation of how vision works at the molecular level.
The experiment revealed the inner workings of an important class of receptors called G-proteincoupled receptors. Rhodopsin is one type of these. G-protein-coupled receptors are present in many systems throughout the body, transmitting a wide variety of chemical signals including hormones and neurotransmitters.
Dysfunctional G-protein-coupled receptors are implicated in diseases that cause vision loss, heart failure, metabolic syndrome, schizophrenia, epilepsy, pain, cancer, fertility disorders and many other conditions. An estimated 50 per cent of current drugs are related to diseases that involve G-protein-coupled receptors.
A clearer understanding of how these receptors efficiently recognize and relay chemical signals broadens our understanding of this major component of human physiology.
Local vibrational coherences drive the primary photochemistry of vision. Philip J. M. Johnson, et al., Nature Chemistry 7, 980–986 (2015).
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