Cloning of a palmitoyl-acyl carrier protein thioesterase from oil palm.
A palmitoyl-acyl carrier protein (ACP) thioesterase cDNA clone was isolated from an oil palm cDNA library. The cDNA was expressed in Escherichia coli as a glutathione S-transferase fusion protein and a crude bacterial extract was assayed for acyl-CoA-hydrolysing activity. The recombinant enzyme was able to hydrolyse medium- and long-chain acyl-CoAs. Northern-blot analysis showed a high level of gene expression in leaf, flower and 15-, 17- and 18-week mesocarp tissues. Low-level gene expression was detected in germinated seedlings and 8- and 12-week mesocarp tissues, but no transcript was detected in any kernel tissues. Southern-blot analysis indicated the presence of a single gene and we have also isolated a genomic clone using the cDNA as a probe. Two genomic fragments were subcloned and a 7 kb contiguous stretch of the oil palm genome was sequenced. Comparison of this sequence with the cDNA sequence identified a putative 93 amino acid transit peptide, most of which is missing from the cDNA. The coding region of the gene consisted of seven exons and six introns.
Othman
A
A
School of Biological Sciences, University of Bristol, Woodland Road, Bristol BS8 1UG, UK. abi.othman@bristol.ac.uk
Lazarus
C
C
Fraser
T
T
Stobart
K
K
D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015698:Genomic Library; D007700:Kinetics; D010938:Plant Oils; D010945:Plants, Edible / Q000201:enzymology* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013869:Thiolester Hydrolases / Q000235:genetics / Q000378:metabolism*
D010938:Plant Oils; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D013869:Thiolester Hydrolases; C018748:oleoyl-(acyl-carrier-protein) hydrolase