*2.1.2 Lysophosphatidic acid acyltransferase*

Coconut oil has 92% saturates and most of its TAGs are trisaturated. Moreover, laurate is found enriched at sn-2 position, which is catalyzed by membrane-bound lysophosphatidic acid acyltransferase (LPAAT) enzyme. Davies et al. detected an enzyme from coconut endosperm, which is a laurate-CoA-preferring LPAAT and active during endosperm maturation [9]. The LPAAT enzyme prefers acyl-CoAs containing C10:0, C12:0, and C14:0 acyl groups as acyl-donor substrates [9]. Knutzon et al. [11] performed the LPAAT protein purification and cloned the corresponding cDNA of this gene from coconut. The gene was then transformed and expressed in *Escherichia coli*, and substrate activity profile of this gene matched that of the coconut enzyme. This copy of LPAAT is the gene named as CCG001603.1 in the first version of coconut genome sequence [5]. Knutzon et al. transformed this gene into a rapeseed transgenic gene line which is expressed of a California bay laurel (*Umbellularia californica*) 12:0-acyl carrier protein thioesterase (BET) and contained up to 50% laurate in its developing seeds [11]. In this transgenic rapeseed with BTE, laurate is found almost exclusively at the *sn-1* and *sn-3* positions of the triacylglycerols. Coexpression of the coconut *LPAAT* gene in the transgenic rapeseeds facilitates efficient laurate deposition at the *sn-2* position and caused the accumulation of trilaurin [11].

Xu et al. cloned the promoter sequence of the LPAAT gene and characterized the promoter by constructing a series of plasmids with promoter sequences with varied length of deletions to promote a β-glucuronidase (GUS) gene. The plasmids were transformed into rice, and the transgenic plants showed that reporter genes with these promoter fragments tend to express specifically in rice endosperm [12]. Yuan et al. transformed *CnLPAAT* into yeast, and tested fatty acid composition indicated that the gene increased the levels of C12:0 and C14:0 in a CnLPAAT-pYES2 transformant [16]. However, heterologous overexpression of CnLPAAT in tobacco (*Nicotiana tabacum* L.) decreased the contents of C12:0 and C14:0 in transgenic tobacco seeds, which could result from low contents of short- and medium-chain FAs (0.22%), which are available in tobacco seeds of the total FAs.

#### *2.1.3 Diacylglycerol acyltransferase*

Besides genes important for MCFA accumulation, there are key genes in TAG biosynthesis pathway that influence oil contents and FA composition. Diacylglycerol acyltransferases (DGAT) and phospholipid:diacylglycerol acyltransferases (PDAT) catalyze diacylglycerol (DAG) to form TAG as the final step in TAG synthesis, using either acyl-CoAs or phospholipids. DAG is an important branch

point between storage and membrane lipid synthesis. Coconut palm has three orthologs of AT2G19450 (*AtDGAT1*) and two orthologs of AT3G51520 (*AtDGAT2*). Coconut *DGATs* genes had higher expression level in coconut endosperm than in the leaf and embryo, especially for *DGAT1* isoform CCG007098.3 and *DGAT2* isoform CCG026159.1 [5].

Zheng et al. cloned a DGAT2 gene from coconut pulp and transferred the gene into the deficient yeast H1246 and *Arabidopsis* [13]. The DGAT2 gene that is expressed in the deficient yeast had DGAT catalysis activity and restored TAG synthesis in the yeast. Further lipid composition analysis showed that *CnDGAT2* has a substrate preference for two UFAs (C16:1 and C18:1) in yeast and linoleic acid (C18:2) in transgenic plants. These results provide knowledge on CnDGAT2 and offer new insights into TAG assembly in coconut.
