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Bioluminescence is the inherent chemical reaction found in certain living organisms that causes things like fireflies to glow and jellyfish to shine. Researchers have been eager to harness these creatures’ light-generating genes to replicate similar effects in vertebrates for various biomedical uses.
Assistant Professor of Biomolecular Engineering at UC Santa Cruz, Andy Yeh, is creating entirely artificial proteins that emit bioluminescence. These proteins aim to provide a non-invasive approach to bioimaging, diagnostic activities, drug discovery, and additional applications. A recent publication in the esteemed journal Chem showcases a new set of bioluminescent proteins developed by Yeh and his team, which are compact, efficient, highly stable, and capable of generating different colors of light for real-time imaging in cellular and animal studies.
This innovative area known as “de novo protein design” recently earned a group of scientists, including David Baker—Yeh’s post-doctoral mentor—the 2024 Nobel Prize in Chemistry. The bioluminescent proteins discussed in this publication were engineered using deep learning software for protein design created by Baker’s research group and protein structure prediction techniques developed by DeepMind, whose founder also shared in the Nobel Prize recognition.
“We refer to this as de novo protein design because these proteins are designed from the ground up, without any natural counterparts or past evolutionary pathways. We demonstrated the use of the recently awarded Nobel Prize concept to create new light-emitting enzymes, which function as optical-based probes for biological research,” Yeh explained.
Enhanced imaging probes surpassing fluorescence
Many scientists and healthcare professionals utilize fluorescence imaging techniques to better understand diseases and support drug discovery, among other purposes. Fluorescence-based probes depend on external light to illuminate. When this light is applied to a tissue, every cell reacts, resulting in excess background light that complicates identifying what researchers or clinicians are focusing on.
In contrast, bioluminescent imaging operates without the need for external excitation—its entire light-emission process occurs at a chemical reaction level. Bioluminescence does not generate background light interference, making it significantly more effective for detecting features that may reside deep within tissues, such as tumors.
This publication confirms that the team’s light-emitting proteins function effectively at the molecular, cellular, and overall animal level, enhancing their applicability across various scientific inquiries. They are particularly well-suited for non-invasive in vivo imaging, as they can provide insights into biological processes deep within tissues in real time without necessitating sample removal from the organism.
Observing multiple biological occurrences
The specially crafted proteins are described as “orthogonal.” Their reaction centers are meticulously designed to fit the constructed light-emitting molecules, ensuring that these designer enzymes do not interact with other similar compounds a researcher or clinician may be utilizing simultaneously.
“The specific nature of the designed reaction allows it to be integrated with existing light-emitting enzymes, as the enzyme identifies a different molecule,” Yeh noted. “Researchers already utilize natural light-emitting enzymes widely in biological studies, and we are not trying to overhaul that. Instead, we are developing additional tools that work effectively and can complement the bioluminescence resources familiar to the scientific community.”
Yeh’s team has also devised a technique for altering the color of the emitted light. Whereas these enzymes traditionally emit blue light, they have enabled the emission of green, yellow, orange, and red through an effective energy transfer process. This advancement permits researchers or clinicians to observe various biological characteristics simultaneously, a method referred to as “multiplexing,” which is essential for investigating intricate processes such as cancer progression.
The dawn of de novo protein design
These de novo proteins demonstrate high thermostability, meaning they will maintain their structure at elevated temperatures, unlike some naturally occurring bioluminescent enzymes. A stable protein would simplify usage in point-of-care diagnostics by eliminating the need for specialized low-temperature shipping.
“We have now synthesized light-emitting enzymes that possess ideal protein folding, something that nature doesn’t always optimize through evolution,” Yeh commented. “This is the first time we’ve shown that artificial light-emitting enzymes can generate enough photons in vertebrate animals for bioimaging purposes. The methods for computational protein design are continuously improving, as will the enzymes we create. I firmly believe in what David Baker has stated: this is merely the beginning of de novo protein design.”
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