PLANT PHYSIOLOGY , Vol 103, Issue 4 1361-1367, Copyright © 1993 by American Society of Plant Biologists
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MOLECULAR BIOLOGY AND GENE REGULATION |
3-Ketoacyl-Acyl Carrier Protein Synthase III from Spinach (Spinacia oleracea) Is Not Similar to Other Condensing Enzymes of Fatty Acid Synthase
H. Tai and J. G. Jaworski
Chemistry Department, Miami University, Oxford, Ohio 45056
A cDNA clone encoding spinach (Spinacia oleracea) 3-ketoacylacyl carrier
protein synthase III (KAS III), which catalyzes the initial condensing
reaction in fatty acid biosynthesis, was isolated. Based on the amino acid
sequence of tryptic digests of purified spinach KAS III, degenerate
polymerase chain reaction (PCR) primers were designed and used to amplify a
612-bp fragment from first-strand cDNA of spinach leaf RNA. A root cDNA
library was probed with the PCR fragment, and a 1920-bp clone was isolated.
Its deduced amino acid sequence matched the sequences of the tryptic
digests obtained from the purified KAS III. Northern analysis confirmed
that it was expressed in both leaf and root. The clone contained a 1218-bp
open reading frame coding for 405 amino acids. The identity of the clone
was confirmed by expression in Escherichia coli BL 21 as a glutathione
S-transferase fusion protein. The deduced amino acid sequence was 48 and
45% identical with the putative KAS III of Porphyra umbilicalis and KAS III
of E. coli, respectively. It also had a strong local homology to the plant
chalcone synthases but had little homology with other KAS isoforms from
plants, bacteria, or animals.
