Researchers show how railroad worms produce red light

September 19, 2019

One research group comprising Brazilian and Japanese scientists have discovered how luciferase produced by the railroad worm Phrixothrix hirtus emits red light.

Luciferase is an enzyme that catalyzes the oxidation of luciferin in fireflies, producing oxyluciferin and enabling fireflies to emit light. Differences in the molecular structures explain the different colors of this bioluminescence in different species.

This discovery has the potential for new biotechnological applications, such as the imaging of muscles, blood and hemoglobin-rich tissue.

An article published in Scientific Reports describes the study, which was conducted by researchers at the Federal University of São Carlos (UFSCar) in São Paulo State, the National Bioscience Laboratory (LNBio) attached to Brazil's National Energy and Materials Research Center (CNPEM), and the University of Electro-Communications in Tokyo, Japan.

The Brazilian team used cloned railroad worm luciferase, which naturally emits red light and mutants of the enzyme together with a larger analog of luciferin synthesized by the Japanese team.

"This novel combination of luciferase with a luciferin analog not only revealed the larger size of the cavity in the luciferase but also produced far-red light more efficiently and is ideal for biomedical applications involving imaging of cells and tissues that preferably absorb blue-green light, such as mammalian cells," Vadim Viviani, a professor at UFSCar (Sorocaba campus) and principal investigator for the study, told.

In a previous study, the group led by Viviani showed that luciferase from fireflies, which are close relatives of railroad worms, changed the color of the light they emitted from green to red in a test tube in response to a change in the acidity of the medium or the presence of heavy metals (read more at:

The group did not yet know how red light was produced naturally by railroad worm luciferase, however, and has now shown how the phenomenon occurs in this species of beetle. Vanessa Rezende Bevilaqua, the first author of the article, participated in the research for her PhD with São Paulo Research Foundation - FAPESP's support.

The study was also part of the FAPESP Thematic Project Arthropod bioluminescence.

P. hirtus is native to the Americas and is one of the few animals known to emit red light as well as green-yellow light, which is more common. During the larval stage, P. hirtus has several green "lanterns" on its back and a red one on its head. The latter helps the beetle find its way in the dark. The light sources on its back serve to frighten predators.

When males of the species become adults, they lose the red lantern but keep the two green ones. Adult females keep them all.

"We've now shown that yellow-green luciferases have a smaller cavity at the active site where luciferin binds and is oxidized to oxyluciferin," Viviani said. "The luciferin is compressed into a more rigid environment, leading to electrostatic repulsion between the two molecules [the energized oxyluciferin and the walls of the luciferase active site], releasing light that contains more energy and is therefore green or yellow."

In the case of the red luciferase produced by the head, the active site cavity is larger, there are more water molecules present, and the environment is less rigid, leading to a reduction in the electrostatic repulsion between the luciferin and the walls of the luciferase active site. This is why the light emitted is red, which contains less energy.


To investigate the interactions that lead to the emission of red light, researchers have learned over the past few decades to clone various luciferases by using genetic engineering tools to modify some amino acids.

The Japanese group, led by Takashi Hirano at the University of Electro-Communications, synthesized red light-emitting luciferin analogs, which were tested by Bevilaqua with the firefly and railroad worm luciferases cloned and modified by the Brazilian research group.

Some of these modified luciferins had a larger structure than the others, and these larger luciferins interacted best with railroad worm luciferase, emitting far-red light more efficiently, whereas they did not interact efficiently with the green or yellow luciferases.

"The luciferases that catalyze green and yellow light have a small cavity and therefore don't bind well to the large-structure luciferin analogs, which have very little luminescent activity," Viviani said. "On the other hand, these large analogs interact well with luciferases that catalyze red light. We deduced from this that railroad worm luciferase has a large active site cavity capable of binding to the analogs."

Having achieved this result, the researchers began testing new combinations of luciferins with modified railroad worm luciferase, eventually creating a more intense red light than that produced by P. hirtus. They believe that the combinations could be used in biomedical research.

"The luciferin analogs synthesized by the Japanese group aren't the first to be created but offer the advantage of having more luminescent activity and a red-shifted light spectrum when they're combined specifically with luciferase from P. hirtus. The commercially available analogs are less efficient, albeit more red-shifted," Viviani said.

Initially, the idea is that this discovery could be used to enhance the visualization of biochemical and cellular processes in mammalian substances that do not absorb red light, such as blood cells and muscle tissue.

"When these substances are examined with conventional luciferase that emits green, yellow or blue light, it's impossible to see biochemical and pathological processes clearly because pigments such as hemoglobin and myoglobin absorb most of the light in these parts of the chromatic spectrum," Viviani said.
About São Paulo Research Foundation (FAPESP)

The São Paulo Research Foundation (FAPESP) is a public institution with the mission of supporting scientific research in all fields of knowledge by awarding scholarships, fellowships and grants to investigators linked with higher education and research institutions in the State of São Paulo, Brazil. FAPESP is aware that the very best research can only be done by working with the best researchers internationally. Therefore, it has established partnerships with funding agencies, higher education, private companies, and research organizations in other countries known for the quality of their research and has been encouraging scientists funded by its grants to further develop their international collaboration. You can learn more about FAPESP at and visit FAPESP news agency at to keep updated with the latest scientific breakthroughs FAPESP helps achieve through its many programs, awards and research centers. You may also subscribe to FAPESP news agency at

Fundação de Amparo à Pesquisa do Estado de São Paulo

Related Molecular Structures Articles from Brightsurf:

Molecular swarm rearranges surface structures atom by atom
Researchers at the University of Münster have now developed a molecular tool which makes it possible, at the atomic level, to change the structure of a metal surface.

UC Davis researchers reveal molecular structures involved in plant respiration
A study published today (Aug. 25, 2020) in eLife provides the first-ever, atomic-level, 3D structure of the largest protein complex (complex I) involved in the plant mitochondrial electron transport chain.

RNA structures by the thousands
Researchers from Bochum and Münster have developed a new method to determine the structures of all RNA molecules in a bacterial cell at once.

Harnessing the domino effect for deployable structures
A team of researchers from the Harvard John A. Paulson School of Engineering and Applied Sciences (SEAS) have harnessed the domino effect to design deployable systems that expand quickly with a small push and are stable and locked into place after deployment.

A new approach to reveal the multiple structures of RNA
The key of the extraordinary functionality of ribonucleic acid, better known as RNA, is a highly flexible and dynamic structure.

How human social structures emerge
What rules shaped humanity's original social networks? The earliest social networks were tightly knit cultural groups made of multiple biologically related families.

Structures in seaweed shed light on sustainability
By examining the enzymes that break down the alginate, the researchers from China and the United Kingdom may be able to harness the natural process to produce biofuel.

A laser, a crystal and molecular structures
Researchers have built a new tool to study molecules using a laser, a crystal and light detectors.

Optimizing structures within complex arrangements of bubbles
New research published in EPJ E explores how different numbers of 2D bubbles of two different areas can be arranged within circular discs, in ways which minimize their perimeters.

A new way of making complex structures in thin films
Self-assembling materials called block copolymers, which are known to form a variety of predictable, regular patterns, can now be made into much more complex patterns that might someday be useful for making optical or plasmonic devices (in which electromagnetic waves interact with electrons), according to an MIT study.

Read More: Molecular Structures News and Molecular Structures Current Events is a participant in the Amazon Services LLC Associates Program, an affiliate advertising program designed to provide a means for sites to earn advertising fees by advertising and linking to