Molecular cloning of starch synthase I from maize (W64) endosperm and expression in Escherichia coli.
A full length cDNA clone encoding a starch synthase (zSS) from maize endosperm (inbred line W64) was isolated and characterized. The cDNA clone (Ss1) is 2907 bp in length and contains an open reading frame of 1866 bp corresponding to a polypeptide of 622 amino acid residues including a transit peptide of 39 amino acids. The Ss1 cDNA clone was identified as zSSI by its direct alignment with sequences to: (i) the N-terminus obtained from the granule-associated form of the zSSI polypeptide, (ii) four internal peptide fragments obtained from the granule-associated form of the zSSI protein, and (iii) one internal fragment from the soluble form of the zSSI protein. The deduced amino acid sequence of Ss1 shares 75.7% sequence identity with rice soluble Ss and contains the highly conserved KSGGLGDV putative ADP-Glc binding site. Moreover, Ss1 exhibited significant activity when expressed in E. coli and the expressed protein is recognized by the antibody raised against the granule associated zSSI protein. Ss1 transcripts were detected in endosperm beginning at 15 days after pollination, but were not found in embryo, leaf or root. Maize contains a single copy of the Ss1 gene, which maps close to the Waxy locus of chromosome 9.
Knight
M E
ME
Zeneca Agrochemicals, Jealotts Hill Research Station, Bracknell, Berkshire, UK.
Harn
C
C
Lilley
C E
CE
Guan
H
H
Singletary
G W
GW
MuForster
C
C
Wasserman
B P
BP
Keeling
P L
PL
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli; D015723:Gene Library; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016366:Open Reading Frames; D012275:Oryza / Q000201:enzymology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D012639:Seeds / Q000201:enzymology; D016415:Sequence Alignment; D013214:Starch Synthase / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D003313:Zea mays / Q000201:enzymology* / Q000235:genetics
D015345:Oligonucleotide Probes; D011994:Recombinant Proteins; D013214:Starch Synthase