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Lin, Myat T.

Publications and source records attributed to Lin, Myat T..

5 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Improving the efficiency of Rubisco by resurrecting its ancestors in the family Solanaceae

Plants and photosynthetic organisms have a remarkably inefficient enzyme named Rubisco that fixes atmospheric CO 2 into organic compounds. Understanding how Rubisco has evolved in response to past climate change is important for attempts to adjust plants to future conditions. In this study, we developed a computational workflow to assemble de novo both large and small subunits of Rubisco enzymes from transcriptomics data. Next, we predicted sequences for ancestral Rubiscos of the (nightshade) family Solanaceae and characterized their kinetics after coexpressing them inEscherichia coli. Predicted ancestors of C 3 Rubiscos were identified that have superior kinetics and excellent potential to help plants adapt to anthropogenic climate change. Our findings also advance understanding of the evolution of Rubisco’s catalytic traits.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Probing photosynthesis by altering chloroplast proteins (Final Technical Report)

The goal of this project was to explore the effect of altering the carbon fixing enzyme Rubisco in the model C3 plant tobacco on photosynthesis and plant growth and development. We introduced the gene sequences encoding Rubisco from a Limonium species and from a red alga into tobacco and found that the enzyme did not assemble properly, undoubtedly requiring additional assembly factors from the source organism. We then developed a method to make single amino-acid changes into the Rubisco subunit that is encoded by the chloroplast genome. We used the method to investigate the role of particular amino acid residues in kinetic properties of Rubisco. Finally, we improved a bacterial expression system that has allowed assembly of active tobacco Rubisco in E. coli. We used the improved system to investigate the kinetic properties of Rubisco enzymes that contain only one type of Rubisco small subunit along with the large subunit.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Escherichia coli amino acid auxotrophic expression host strains for investigating protein structure–function relationships

Abstract A set of C43(DE3) and BL21(DE3) Escherichia coli host strains that are auxotrophic for various amino acids is briefly reviewed. These strains require the addition of a defined set of one or more amino acids in the growth medium, and have been specifically designed for overproduction of membrane or water-soluble proteins selectively labelled with stable isotopes, such as 2H, 13C and 15N. The strains described here are available for use and have been deposited into public strain banks. Although they cannot fully eliminate the possibility of isotope dilution and mixing, metabolic scrambling of the different amino acid types can be minimized through a careful consideration of the bacterial metabolic pathways. The use of a suitable auxotrophic expression host strain with an appropriately isotopically labelled growth medium ensures high levels of isotope labelling efficiency as well as selectivity for providing deeper insight into protein structure–function relationships.

Biochemistry & Molecular Biology↗
DOE OSTI2020

Small subunits can determine enzyme kinetics of tobacco Rubisco expressed in Escherichia coli

Rubisco catalyses the first step in carbon fixation and is a strategic target to improve photosynthetic efficiency. In plants, Rubisco is composed of eight large and eight small subunits and its biogenesis requires multiple chaperones. We optimised a system to produce tobacco Rubisco in Escherichia coli by co-expressing chaperones in auto-induction medium. We successfully assembled tobacco Rubisco in E. coli with each small subunit that is normally encoded by the nuclear genome. Even though each enzyme carries only a single type of small subunit in E. coli, the enzymes exhibit carboxylation kinetics very similar to that of the native Rubisco. Tobacco Rubisco assembled with a recently discovered trichome small subunit has a higher catalytic rate and a lower CO 2 affinity than those assembled with other small subunits. Finally, our E. coli expression system will allow probing of features of both subunits of Rubisco that affect its kinetic properties.

59 BASIC BIOLOGICAL SCIENCES↗