ISOLATION AND CHARACTERIZATION OF A MAIZE CDNA THAT COMPLEMENTS A 1-ACYL SN-GLYCEROL-3-PHOSPHATE ACYLTRANSFERASE MUTANT OF ESCHERICHIA-COLIAND ENCODES A PROTEIN WHICH HAS SIMILARITIES TO OTHER ACYLTRANSFERASES
Ap. Brown et al., ISOLATION AND CHARACTERIZATION OF A MAIZE CDNA THAT COMPLEMENTS A 1-ACYL SN-GLYCEROL-3-PHOSPHATE ACYLTRANSFERASE MUTANT OF ESCHERICHIA-COLIAND ENCODES A PROTEIN WHICH HAS SIMILARITIES TO OTHER ACYLTRANSFERASES, Plant molecular biology, 26(1), 1994, pp. 211-223
We selected cDNA plasmid clones that corrected the temperature-sensiti
ve phenotype of Escherichia coli strain JC201, which is deficient in 1
-acyl-sn-glycerol-3-phosphate acyltransferase activity. A plasmid-base
d maize endosperm cDNA library was used for complementation and a plas
mid that enabled the cells to grow at 44 degrees C on ampicillin was i
solated. Addition of this plasmid (pMAT1) to JC201 restored 1-acyl-sn-
glycerol-3-phosphate acyltransferase activity to the cells. Total phos
pholipid labelling showed that the substrate for the enzyme, lysophosp
hatidic acid, accumulated in JC201 and was further metabolised to phos
phatidylethanolamine in complemented cells. Membranes isolated from su
ch cells were able to convert lysophosphatidic acid to phosphatidic ac
id in acyltransferase assays. The cDNA insert of pMAT1 contains one lo
ng open reading frame of 374 amino acids which encodes a protein of re
lative molecular weight 42 543. The sequence of this protein is most s
imilar to SLC1, which is thought to be able to acylate glycerol at the
sn-2 position during synthesis of inositol-containing lipids. Homolog
ies between the SLC I protein, the 1-acyl-sn-glycerol-3-phosphate acyl
transferase of E. coli (PlsC) and the maize ORF were found with blocks
of conserved amino acids, whose spacing was conserved between the thr
ee proteins, identifiable.