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Title:	Characterisation And Expression Of Oil Palm (Elaeis Guineensis) GDSL And GXSXG Lipase Genes
Authors:	Wong Yun Teng (P46458)
Supervisor:	Wickneswari Ratnam, Prof. Dr.
Keywords:	Characterisation Of Oil Palm Expression Of Oil Palm GDSL Lipase Genes GXSXG Lipase Genes Oil palm
Issue Date:	4-Jul-2013
Description:	Lipases have been recognised as very useful biocatalysts due to their great versatility in industrial applications. However, not much is known about plant lipases as compared to microbial and animal lipases. This is mainly due to their low abundance and difficulty in purification. Previous work confirmed the existence of an endogenous lipase in Elaeis guineensis (oil palm) mesocarp and this enzyme is activated at low temperature. Therefore, in this study, we sought to identify and characterise the lipase gene, as well as to investigate the expression profile of the gene. In this study, a unique lipase gene containing a GDSL motif and belong to the SGNH superfamily was identified in the oil palm. Prior to this, the oil palm was found to contain the commonly occurring GXSXG lipase gene. Primers were designed based on a partial cDNA sequence (360bp) from expressed sequence tags (ESTs) was used to isolate the full length cDNA, using a Rapid Amplification of cDNA Ends Polymerase Chain Reaction (RACE PCR). A full length cDNA clone with an open reading frame (ORF) of 1083bp encoding 360 amino acid residues was identified. The DNA and protein sequence of this lipase gene was obtained and characterized. The amino acid sequences was compared using BLAST; primary structure analysis was performed using ExPASY and Protscale and a multiple sequence alignment was done using the ClustalW algorithm. A three-dimensional model for this protein was predicted and built using 3D-JIGSAW, based on homologues of the known protein sequence. This GDSL lipase was confirmed to have a catalytic triad (Ser32, Asp332 and His335), as well as oxyanion hole residues (Ser32, Gly64, or Gly69, Asn167) and five other highly conserved peptide blocks. Thus, it is similar to other carboxylester hydrolases of this family. Southern blotting implicated a single or low copy number of lipase gene in oil palm. Screening of oil palm germplasm collection confirmed previous studies that the Angolan accession had highest and Guinea accession the lowest lipase activity based on free fatty acid (FFA) content in the mesocarp. Real-time PCR showed significant positive Pearson correlation of r = 0.397 and 0.194 respectively of GDSL and GXSXG gene expression with lipase activity in the oil palm mesocarp. This suggests that the GDSL lipase plays a larger role than GXSXG in FFA increases in the mesocarp. However, the expression of GDSL and GXSXG are highly correlated (r = 0.872) suggesting that the two enzyme may work in tandem. In addition, the expression studies showed that this GDSL lipase is active in the oil palm mesocarp, leaf and most active in roots in contrast to the GXSXG lipase which is only expressed in the mesocarp. These findings suggest GDSL lipase has a broader substrates and functions as compared to the common lipase in the oil palm.,PhD
Pages:	156
Call Number:	QK495.P17 .W646 2013
Publisher:	UKM, Bangi
Appears in Collections:	Faculty of Science and Technology / Fakulti Sains dan Teknologi

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