Constitution of stable artificial oil bodies with triacylglycerol, phospholipid, and caleosin

Seed oil bodies are lipid storage organelles of 0.5-2 microm in diameter and comprise a triacylglycerol matrix shielded by a monolayer of phospholipids and proteins. These proteins include abundant structural proteins, oleosins, and at least two minor proteins termed caleosin and steroleosin. This study examined if artificial oil bodies (AOBs) composed of triacylglycerol and phospholipid could be stabilized by oleosin, caleosin, or steroleosin. Our results showed that stabilization effects could be realized by oleosin or caleosin but not by steroleosin. The sizes of the AOBs constituted with oleosin (0.5-2 microm) or caleosin (50-200 nm) were similar to or 10 times smaller than those of the native oil bodies. Recombinant caleosin expressed in Escherichia coli also encapsulated AOBs with a size, topology, and stability comparable to those encapsulated with native caleosin. A proteinase K digestion indicated that caleosin anchored the AOBs via its central hydrophobic domain of approximately 4 kDa. Isoelectrofocusing revealed that the isoelectric point of the caleosin-stabilized AOBs was pH 4.0. Aggregation of AOBs was observed at a pH lower than 4.5; thus, their stability and integrity were presumably contributed by surface caleosin via electronegative repulsion and steric hindrance. The caleosin-stabilized AOBs were thermostable up to 70 degrees C and potentially useful for biotechnological applications.