The Philippine Agricultural Scientist Vol. 91 No. 4 (December 2008) 389
Oil Bodies and Oleosin of Coconut A. J. D. Rodelas et al.
THE PHILIPPINE AGRICULTURAL SCIENTIST ISSN 0031-7454
Vol. 91 No. 4, 389-394 December 2008
Isolation and Characterization of the Oil Bodies and Oleosin of Coconut (Cocos nucifera L.)
Abigail Joy D. Rodelas
§, Ellen S. Regalado
§, Dennis B. Bela-ong, Roberta N. Garcia, Antonio C. Laurena and Evelyn Mae Tecson Mendoza
*Institute of Plant Breeding, Crop Science Cluster, College of Agriculture, University of the Philippines Los Baños, College, Laguna 4031, Philippines
§AJDR and ESR contributed equally to this paper.
*Author for correspondence; e-mail: [email protected], [email protected]
Oil bodies were isolated from the solid endosperm of 11–12 month old coconut and purified by sucrose density gradient washing, urea washing and floatation centrifugation. Under light microscope, purified oil bodies showed two types of small and large oil bodies: about 90% with diameter of 0.5–4.0 µµµµµ
µµµµ
m and 10%, 10–40 µm, respectively. The oil bodies consisted of 97% triacylglycerols, phospholipids (phosphatidylethanoloamine and phosphatidylcholine) and the oil body storage protein oleosin. Puri- fied oleosin showed one band of 14400 on SDS-PAGE. Two-dimensional electrophoresis resolved one band of 14400 with an isoelectric pH between 9 and 10. The 14400 band had an N-terminal amino acid sequence of GEERR or GEEER. Additional urea washings of the coconut oil bodies followed by ether delipidation revealed two oleosin bands of 14400 and 11000 Mr on SDS-PAGE.
Key Words: oleosin, oil body, coconut, Cocos nucifera, oil body storage protein
INTRODUCTION
Oil bodies are small spherical organelles, usually ranging from 0.4–3 mm in diameter (Murphy and Cummins 1989;
Tzen et al. 1993), which are deposition sites of plant stor- age lipids (Huang 1992) and are composed mainly of triacylglycerols (TAGs) covered by a protein-phospho- lipid complex. They are synthesized during the early to middle stages of seed development (Murphy et al. 1989).
Their size may vary depending on the source species, cy- toplasmic composition and viscosity (Sarmiento et al.
1997), as well as nutrition, environment, oil to oleosin ratio and the organism’s genetic background (Ting et al. 1996).
They are used as energy reserves for germination and are immobilized during prolonged storage (Li et al. 2002). Larger oil bodies of 10–20 µm have been observed in the fleshy mesocarp of olive drupes (Ross et al. 1993) and avocado (Platt-Aloia and Thomson 1981).
Oleosins are a unique class of proteins found in the oil bodies of diverse plant species. They are embedded in the phospholipid coat of oil bodies and are responsible for the oil body size and stability (Tzen et al. 1997). They are small, largely hydrophobic proteins ranging in size from 15–26 kDa (Huang 1992) and are composed of three re-
gions: an N-terminal hydrophilic region of variable length (from 6–60 residues), a central highly conserved hydro- phobic domain of 72 residues and a C-terminal amphipathic region of variable length, from 28 to as long as 1000 resi- dues in Arabidopsis oleosins (Hsieh and Huang 2004). The central hydrophobic domain, a distinctive trait of oleosins, forms a hairpin with two arms of 30 amino acids joined by a head called the proline knot (Hsieh and Huang 2004).
Oleosins are most abundant in the lipid-storing bodies of seeds comprising up to 2–8% of the total seed proteins although they have also been found in various parts of plants such as the tapetum cells of anthers (Hsieh and Huang 2004; de Oliveira et al. 1993; Roberts et al. 1994) in the Brassicaceae family. Oleosins were observed only in the smaller oil bodies (0.5–2.0 µm) in the embryo and en- dosperm tissues of the olive seed (Ross et al. 1993).
Researchers have suggested different roles for oleosins such as in the stabilization of oil bodies prevent- ing their aggregation (Tzen and Huang 1992) and as recep- tor for the binding of lipase to mobilize triacylglycerols during germination and post-germinative growth.
Coconut (Cocos nucifera L.) is a monospecies plant belonging to the Arecaceae family (Harries 1978). The solid endosperm of its seed consists of about 45–55% oil, which
