Computational Prediction of Mutagenesis in Glycine max Rubisco Activase Monomer for Increased Thermal Stability

Increasing global temperatures compromise the rate of photosynthesis in plants. Although several factors contribute, the enzyme Rubisco Activase (RCA) has been identified as a key limitation to photosynthesis at temperatures above 35^oC. RCA activates the carbon fixation enzyme Rubisco but has low thermal stability. We computationally predicted single point mutations that would increase thermostability of RCAβ from soybean (Glycine max). We compared several existing computational tools and found that PremPS most closely matched existing data and literature values for this protein. A set of 456 mutations were tested with 32.33% predicted to stabilize the protein structure of soybean RCA. The three most stabilizing mutants were expressed in E. coli and characterized by in vitro ATPase activity at a range of temperatures. All three computationally predicted mutations significantly increased the thermostability compared to the wild-type protein.