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GENOMICS, MOLECULAR GENETICS & BIOTECHNOLOGY

Sulfur Assimilation in Soybean

Molecular Cloning and Characterization of O-Acetylserine (Thiol) Lyase (Cysteine Synthase)

^a Dep. of Agronomy, Univ. of Missouri, Columbia, MO 65211
^b USDA-ARS, Plant Genetics Research Unit, Univ. of Missouri, Columbia, MO 65211

^* Corresponding author (KrishnanH{at}missouri.edu).

Soybean [Glycine max (L.) Merr.] is a good protein source for both humans and livestock. However, soybean seed proteins are deficient in the sulfur-containing amino acids cysteine and methionine. This deficiency has stimulated efforts to improve the amino acid composition of soybean seed proteins. Our overall goal is to improve the sulfur amino acid content of soybean seed proteins by genetic manipulation. The objective of this study was to isolate and characterize O-acetylserine (thiol) lyase (OAS-TL), a key enzyme that catalyzes the last step in the production of cysteine. A full-length cDNA clone encoding a cytosolic isoform of OAS-TL was isolated by screening a soybean seed cDNA library with a ³²P-labeled expressed sequence tag (EST). Nucleotide sequence analysis of the cDNA revealed a single open-reading frame of 978 base pairs (bp) encoding a 34-kDa protein. The authenticity of the isolated cDNA was confirmed by the functional complementation of an Escherichia coli cysteine auxotrophic mutant. Reverse transcriptase polymerase chain reaction (RT-PCR) analysis revealed that OAS-TL mRNA was abundant at early stages of seed development. Western blot analysis using antibodies generated against the recombinant soybean OAS-TL demonstrated that the abundance of this protein gradually declined during later stages of seed development. The OAS-TL activity peaked in young developing seeds and declined steadily during the time period when the bulk of seed storage protein accumulation occurred. Thus, elevating the specific activity of OAS-TL during later stages of seed development could lead to an increase in cysteine synthesis in soybean seeds.

Abbreviations: OAS-TL, O-acetylserine (thiol) lyase • ORF, open reading frame • PCR, polymerase chain reaction • RT-PCR, reverse transcriptase polymerase chain reaction • SAT, serine acetyltransferase

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				H. B. Krishnan Engineering Soybean for Enhanced Sulfur Amino Acid Content Crop Sci., January 31, 2005; 45(2): 454 - 461. [Abstract] [Full Text] [PDF]