PNNL

RICHLAND, Wash.—Traditionally, biotechnology researchers have modified genes when engineering microbes for more robust and efficient production of biofuels and bioproducts. But researchers at Pacific Northwest National Laboratory (PNNL) are using predictive phenomics to uncover additional layers of biological control, tracking how environmental changes reshape molecular activity inside a cell and how those shifts translate to function. 

The team focused on photosynthetic microbes like Synechococcus—better known as blue‑green algae—that have evolved to respond rapidly to changes in light. Those responses can quickly alter how the microbes harvest light, store energy, and fix carbon—central processes in both natural ecosystems and industrial bioproduction.

Understanding how these bacteria work is important to unlocking new potential for biotechnology solutions, where microbes play important roles in industrial processes to make chemicals, energy, fuels, and biomaterials. Last year, in a paper in Microbial Cell Factories, a PNNL research team led by Pavlo Bohutskyi showed how disrupting the circadian rhythm of Synechococcus elongatus PCC 7942 could dramatically boost the bacteria’s output—a step toward producing biology‑based products more efficiently and less expensively. 

That earlier study focused primarily on gene expression. But gene activity alone doesn’t fully explain how cells behave as conditions change, a central motivation behind PNNL’s Predictive Phenomics Initiative. “If we want to engineer cyanobacteria for efficient industrial bioproduction, we can’t just focus on manipulating genes,” said Bohutskyi. “There is a hidden layer of protein regulation where the organism fine‑tunes its output.”

In a new study published in Molecular & Cellular Proteomics, a team led by John Melchior looked directly at that protein layer. They measured not just which proteins increase or decrease in abundance, but how the proteins change shape, stability, and chemical state as light conditions shift.

To capture those dimensions of change, the team combined traditional proteomics with three complementary “structural proteomics” methods. Limited proteolysis mass spectrometry (LiP‑MS) detects proteins that change shape. Thermal proteome profiling (TPP‑MS) measures changes in protein stability that can reflect new interactions with partner proteins or metabolites. Redox proteomics tracks rapid chemical modifications—often involving cysteine residues—that can act as fast regulatory switches during stress and signaling. Their results provide deeper insights into microbial adaptability and potential engineering targets for biotechnology applications.

“You can think of it this way: Traditional proteomics shows us who’s in the room,” said Melchior. “The structural changes and stability measurements show us what they’re doing, who may be interacting, and how their roles change when the lights shift. Without all of those tools together, the story is incomplete.” 

The combined techniques identified previously hidden protein dynamics and highlight why genomics and transcript measurements by themselves can miss important control points. By pinpointing protein-level regulation that drives rapid shifts in photosynthesis and metabolism, the work may help guide efforts to design strains that are resilient and adaptable in stressful conditions, necessary for bioproduction.

The researchers found that many of the most important responses happen through rapid protein remodeling, not simply by making more or less of a protein. In a transient dark treatment, only 145 proteins changed in abundance, but more than 400 showed structural changes. When cells were shifted from light‑limited to intense light for 30 minutes, abundance measurements captured changes in 904 proteins, while structural measurements revealed an additional 3,021 structural or chemical changes across proteins.

In the current study, rapid changes were concentrated in core systems that control how cyanobacteria capture light and route energy, including the light‑harvesting phycobilisome antenna and photosystems, the electron transport chain, ribosomes that regulate protein synthesis, and central carbon metabolism pathways involved in carbon fixation and energy storage. For example, the team detected coordinated structural and redox changes in cytochrome f (petA), a key component that controls electron flow between the photosystems, that were not apparent from abundance measurements alone.

Their findings highlight how phenomics can open new avenues for biotechnology solutions by improving predictive models of how cells reprogram themselves under variable conditions relevant to biomanufacturing. While scientists have generally focused on modifying genes, this study highlights additional properties—rapid changes in protein shape, stability and redox state—that could be leveraged to build bacterial strains that stay productive under fluctuating light and other real-world conditions. 

The work is part of a research effort called the Predictive Phenomics Initiative at PNNL. Researchers are studying how changes in the environment affect the molecular processes in an organism and how these affected processes change an organism’s functions. The team plans to use the findings from the current study to build a predictive understanding of how Synechococcus responds to environmental changes and to identify protein‑level engineering targets for industrial biotechnology applications.