Overexpression of ATP sulfurylase improves the sulfur amino acid content, enhances the accumulation of Bowman-Birk protease inhibitor and suppresses the accumulation of the b-subunit of b-conglycinin in soybean seeds

Authors: KIM, WON-SEOK - University Of Missouri; Kim, Sunhyung; Oehrle, Nathan; JEZ, JOSEPH - Washington University; Krishnan, Hari

Submitted to: Scientific Reports
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/26/2020
Publication Date: 9/14/2020
Citation: Kim, W., Kim, S.N., Oehrle, N.W., Jez, J.M., Krishnan, H.B. 2020. Overexpression of ATP sulfurylase improves the sulfur amino acid content, enhances the accumulation of Bowman-Birk protease inhibitor and suppresses the accumulation of the b-subunit of b-conglycinin in soybean seeds. Scientific Reports. 10. Article e14989. https://doi.org/10.1038/s41598-020-72134-z.
DOI: https://doi.org/10.1038/s41598-020-72134-z

Interpretive Summary: Methionine is considered as an “essential” amino acid because humans and monogastric animals cannot synthesize this metabolite and are dependent on obtaining it through food /feed that is being consumed. Though soybeans are an excellent protein source, its quality could be further enhanced if the methionine content of soybeans could be elevated. Biotechnological approaches have been undertaken to elevate the methionine content of soybean seeds. However, the overall improvement of the methionine content has not been satisfactory to meet the poultry and livestock requirements. In this study, we have we generated transgenic soybean plants that overexpress ATP sulfurylase, a key enzyme involved in the sulfur assimilatory pathway. Soybean seeds overexpressing ATP sulfurylase revealed altered seed protein composition and more importantly the sulfur amino acid content seeds were significantly improved. The results obtained from this study will benefit the US soybean farmers by providing tools that will aid in the development of soybeans with higher nutritive value.

Technical Abstract: ATP sulfurylase, an enzyme which catalyzes the conversion of sulfate to adenosine 5'-phosphosulfate(APS), plays a significant role in controlling sulfur metabolism in plants. In this study, we have expressed soybean plastid ATP sulfurylase isoform 1 in transgenic soybean without its transit peptide under the control of the 35S CaMV promoter. Subcellular fractionation and immunoblot analysis revealed that ATP sulfurylase isoform 1 was predominantly expressed in the cell cytoplasm. Compared with that of untransformed plants, the ATP sulfurylase activity was about 2.5-fold higher in developing seeds. High-resolution 2-D gel electrophoresis and immunoblot analyses revealed that transgenic soybean seeds overexpressing ATP sulfurylase accumulated very low levels of the ß-subunit of ß-conglycinin. In contrast, the accumulation of the cysteine-rich Bowman–Birk protease inhibitor was several fold higher in transgenic soybean plants when compared to the non-transgenic wild-type seeds. The overall protein content of the transgenic seeds was lowered by about 3% when compared to the wild-type seeds. Metabolite profiling by LC–MS and GC–MS quantified 124 seed metabolites out of which 84 were present in higher amounts and 40 were present in lower amounts in ATP sulfurylase overexpressing seeds compared to the wild-type seeds. Sulfate, cysteine, and some sulfur-containing secondary metabolites accumulated in higher amounts in ATP sulfurylase transgenic seeds. Additionally, ATP sulfurylase overexpressing seeds contained significantly higher amounts of phospholipids, lysophospholipids, diacylglycerols, sterols, and sulfolipids. Importantly, over expression of ATP sulfurylase resulted in 37–52% and 15–19% increases in the protein-bound cysteine and methionine content of transgenic seeds, respectively. Our results demonstrate that manipulating the expression levels of key sulfur assimilatory enzymes could be exploited to improve the nutritive value of soybean seeds.