Supplying food for a growing world population and drawing down atmospheric CO2 are major 21st Century challenges. Yield increase by conventional breeding is expected to plateau; however, production must increase by ~50% by 2050 to meet demand. Here, synthetic biology and metabolic engineering can offer solutions. One promising but unexplored strategy is to decrease respiratory carbon losses. Part of plant respiration fuels protein turnover (degradation and resynthesis), which in total consumes 10-15% of photosynthetically fixed carbon. Thus, cutting protein turnover costs is a prime crop improvement target. The carbon saved thereby could be channeled into yield or recalcitrant polymers to sequester atmospheric carbon.
Various abundant, short-lived enzymes are candidates for cutting turnover rates. One example is methionine synthase (MetSyn), whose life is likely shortened by damage done to the active site by the reactive homocysteine substrate and folate cofactor. We evolved Arabidopsis MetSyn 1 and 2 towards longer lifespan in the yeast OrthoRep continuous directed evolution system. Putatively beneficial mutant sequences are currently being validated in growth and biochemical assays. Another example is THI4 thiazole synthase, a suicide enzyme (i.e., performs only one catalytic cycle) in which an active-site Cys is destroyed to provide the sulfur atom for the product. The inactivated enzyme must then be replaced. Arabidopsis and barley THI4s turn over faster than any other protein measured, the metabolic cost of this turnover being equivalent to as much as 2-4% of biomass. We are seeking to lengthen the lifespan of (i) three unusual cereal THI4s that have no active-site Cys and appear to be catalytic but very inefficient, and (ii) Arabidopsis THI4 mutants whose active-site Cys is replaced by His, Glu, or Arg. When substituted for the native Arabidopsis THI4, the evolved enzymes are predicted to turn over more slowly, thus reducing respiration and increasing biomass accumulation.