|RuBisCo catalyzes the rate-limiting step in biological carbon dioxide fixation. Source: Emory University|
In research recently completed at Emory University School of Medicine, scientists have discovered a mutant enzyme that could enable plants to use and to convert carbon dioxide more quickly, effectively removing more greenhouse gasses from the atmosphere.
During photosynthesis, plants—and some bacteria—rely on the enzyme rubulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO) to catalyze the rate-limiting step of the Calvin Cycle, the major pathway of carbon dioxide fixation. In other words, RuBisCO controls the rate at which plants can take up carbon dioxide from the atmosphere.
While RuBisCO is the most abundant enzyme in the world, it is also one of the least efficient.
All life pretty much depends on the function on this enzyme. It actually has had billions of years to improve, but remains about a thousand times slower than most other enzymes. Plants have to make tons of it just to stay alive.—Dr. Ichiro Matsumura, senior author and principal investigator
For decades, scientists have struggled to engineer a variant of the enzyme that would more quickly convert carbon dioxide. Their attempts primarily focused on mutating specific amino acids within RuBisCO, and then seeing if the change affected carbon dioxide conversion. Because of RuBisCO’s structural complexity, the mutations did not have the desired outcome.
Previous attempts to improve the catalytic efficiency and/or CO2 specificity of RuBisCO by structure-based site-directed mutagenesis have at best led to modest (5–13%) improvements in specificity, according to the Emory team.
For the Emory study, Dr. Matsumura and his colleagues used directed evolution—isolating and randomly mutating genes, and then inserting the mutated genes into bacteria (in this case Escherichia coli, or E. coli). They then screened the resulting mutant proteins for the fastest and most efficient enzymes.
Because E. coli does not normally participate in photosynthesis or carbon dioxide conversion, it does not usually carry the RuBisCO enzyme. The scientists withheld other nutrients from the genetically modified bacteria so that it would need RuBisCO and carbon dioxide to survive under these stringent conditions.
The fastest growing strains carried mutated RuBisCO genes that produced a larger quantity of the enzyme, leading to faster assimilation of carbon dioxide gas. The RuBisCO variants that evolved during three rounds of random mutagenesis and selection were over-expressed, and exhibited 5-fold improvement in specific activity relative to the wild-type enzyme.
These mutations caused a 500 percent increase in RuBisCO expression. We are excited because such large changes could potentially lead to faster plant growth. This result also suggests that the enzyme is evolving in our laboratory in the same way that it did in nature.—Dr. Matsumura
The findings were published online and in the February issue of the journal Protein Engineering Design and Selection.
“Directed evolution of RuBisCO hypermorphs through genetic selection in engineered E. coli” Monal R. Parikh, Dina N. Greene, Kristen K. Woods, and Ichiro Matsumura; Protein Engineering Design and Selection, doi:10.1093/protein/gzj010