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PLANT PHYSIOLOGY , Vol 103, Issue 2 565-573, Copyright © 1993 by American Society of Plant Biologists
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MOLECULAR BIOLOGY AND GENE REGULATION |
Identification, cDNA Cloning, and Gene Expression of Soluble Starch Synthase in Rice (Oryza sativa L.) Immature Seeds
T. Baba, M. Nishihara, K. Mizuno, T. Kawasaki, H. Shimada, E. Kobayashi, S. Ohnishi, Ki. Tanaka and Y. Arai
Institute of Applied Biochemistry, University of Tsukuba (T.B., M.N., K.M., E.K., S.O.,K.-i.T., Y.A.), and Mitsui Plant Biotechnology Research Institute, (T.K., H.S.), Sengen 2-1-6, Tsukuba Science City, Ibaraki 305, Japan
Three forms of soluble starch synthase were resolved by anion-exchange
chromatography of soluble extracts from immature rice (Oryza sativa L.)
seeds, and each of these forms was further purified by affinity
chromatography. The 55-, 57-, and 57-kD proteins in the three preparations
were identified as candidates for soluble starch synthase by western blot
analysis using an antiserum against rice granule-bound starch synthase. It
is interesting that the amino-terminal amino acid sequence was identical
among the three proteins, except that the 55-kD protein lacked eight amino
acids at the amino terminus. Thus, these three proteins are products of the
same gene. The cDNA clones coding for this protein have been isolated from
an immature rice seed library in [lambda]gt11 using synthetic
oligonucleotides as probes. The deduced amino acid sequence of this protein
contains a lysine-X-glycine-glycine consensus sequence for the
ADP-glucose-binding site of starch and glycogen synthases. Therefore, we
conclude that this protein corresponds to a form of soluble starch synthase
in immature rice seeds. The precursor of the enzyme contains 626 amino
acids, including a 113-residue transit peptide at the amino terminus. The
mature form of soluble starch synthase shares a significant but low
sequence identity with rice granule-bound starch synthase and Escherichia
coli glycogen synthase. However, several regions, including the
substrate-binding site, are highly conserved among these three enzymes.
Blot hybridization analysis demonstrates that the gene encoding soluble
starch synthase is a single-copy gene in the rice genome and is expressed
in both leaves and immature seeds. These results suggest that soluble and
granule-bound starch synthases play distinct roles in starch biosynthesis
of plant.
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