Photosynthesis feeds the world, but it’s not as efficient as it could be. The enzyme Rubisco, responsible for capturing carbon dioxide from the air, is notoriously slow and often struggles to distinguish between carbon dioxide and oxygen, creating a major drag on crop productivity. Scientists have long sought to improve Rubisco, but testing new versions of the enzyme in living plants has been very difficult—the native enzyme gets in the way.
In a new study published in PNAS, our lab solved this problem by creating a tobacco platform that completely lacks native Rubisco. Using CRISPR-Cas9, the team knocked out all 11 genes encoding the small subunit of Rubisco, while keeping plants alive with a backup cyanobacterial version of the enzyme expressed in the chloroplast.
This Rubisco-null platform provides a “clean slate,” allowing us to test new enzyme variants in a living plant without any interference from native proteins. We used this platform to test something remarkable: ancestral Rubiscos. These are “resurrected” ancestors of the Rubisco that date back to eras with higher temperatures and carbon dioxide levels. Our lab previously resurrected these enzymes in E. coli system and found them to be functional and kinetically superior. With the plant system developed in this study, we were able to test these ancestral rubiscos in planta, without native rubiscos muddying the analysis. These “resurrected” ancestral enzymes weren’t just functional—they were faster and more efficient than modern versions, showing 16-20% higher catalytic efficiency under normal atmospheric conditions.
The key to their superior performance? These ancient enzymes work faster, processing carbon dioxide at a significantly higher rate than their modern descendants and are better adapted for higher temperatures and ambient carbon dioxide. The transgenic plants expressing these ancestral Rubiscos appeared completely normal and produced enzyme levels comparable to those of regular tobacco plants.
Improving Rubisco is a “holy grail” of plant science. A more efficient Rubisco means crops could potentially grow faster with less nitrogen and better water-use efficiency. This work establishes ancestral reconstruction as a promising strategy for finding better Rubiscos and provides researchers with a powerful new tool for testing photosynthetic improvements. With crop yields struggling to keep pace with global food demands, unlocking Rubisco’s potential could be crucial for feeding the future.