Bibliography
Cloning, characterization and developmental expression of the coconut (Cocos nucifera L.) 11S globulin gene [2010]
Caldo, K. M. P., Philippines Univ. Los Baños, College, Laguna (Philippines);
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Cocosin, the 11S globulin of coconut, is the major storage protein that accumulates in the endosperm during seed development. This work describes the gene structure, derived amino acid sequence, structural homologies, and developmental expression profile of cocosin. Two full-length cocosin cDNA isoforms were isolated through the PCR cloning strategy coupled with 5' and 3' - RACE technologies. The open reading frames of CnCos-1 and CnCos-2 both span 1,401 nucleotides, which code for 466 amino acid residues. The two isoforms are 92.3 and 91.5% identical in terms of cDNA and amino acid sequences, respectively. The derived amino acid sequences of cocosin exhibit significant homology with 11S globulins/glutelins of various seed plants. The first 30 residues of CnCos-1 and CnCos-2 correspond to a single peptide responsible for targeting cocosin into storage vacuoles. The highly conserved asparaginyl splice site is present in both isoforms and divides the polypeptides into acidic (32 kDa) and basic (21 kDa) subunits. The highly conserved bicupin domain and four cysteine residues involved in disulfide bonding among 11S globulins were mapped in both isoforms. Cocosin is methionine-rich as compared to most angiosperm legumins. Cocosin is not a potential allergen as it does not exhibit substantial identity with any immunodominant epitopes of 11 S globulins implicated in food allergies. Phylogenetic analysis showed that cocosin is more closely related to Elaeis guinee
nsis glutelin, both of which form a distinct clade in the tree between the divergent clades of dicot and cereal 11S globulins. Relative PCR using actin as internal control showed that the synthesis of cocosin transcripts started at 6-7 months after pollination (MAP). The level of cocosin transcripts increased dramatically at 7-8 MAP, and slightly increased further at 8-9 MAP. Polyclonal antibodies against cocosin were raised in white rabbits and used as probes in the immunodetection of cocosin. The antibodies were shown to be highly specific to cocosin through the Ouchterlony Assay and ELISA. Western blot analysis showed that approximately 4.5% of total cocosin was present at 6-7 months after pollination (MAP) and this increased gradually, reaching 45% at 10-11 MAP. The immunoblot further showed that majority of cocosin (approximately 55%) were deposited between the 10-11 and 11-12 MAP.
