Cloning and secondary structure analysis of caleosin, a unique calcium-binding protein in oil bodies of plant seeds.
Plant seed oil bodies comprise a matrix of triacylglycerols surrounded by a monolayer of phospholipids embedded with abundant oleosins and some minor proteins. Three minor proteins, temporarily termed Sops 1-3, have been identified in sesame oil bodies. A cDNA sequence of Sop1 was obtained by PCR cloning using degenerate primers derived from two partial amino acid sequences, and subsequently confirmed via immunological recognition of its over-expressed protein in Escherichia coli. Alignment with four published homologous sequences suggests Sop1 as a putative calcium-binding protein. Immunological cross-recognition implies that this protein, tentatively named caleosin, exists in diverse seed oil bodies. Caleosin migrated faster in SDS-PAGE when incubated with Ca2+. A single copy of caleosin gene was found in sesame genome based on Southern hybridization. Northern hybridization revealed that both caleosin and oleosin genes were concurrently transcribed in maturing seeds where oil bodies are actively assembled. Hydropathy plot and secondary structure analysis suggest that caleosin comprises three structural domains, i.e., an N-terminal hydrophilic calcium-binding domain, a central hydrophobic anchoring domain, and a C-terminal hydrophilic phosphorylation domain. Compared with oleosin, a conserved proline knot-like motif is located in the central hydrophobic domain of caleosin and assumed to involve in protein assembly onto oil bodies.
Chen
J C
JC
Graduate Institute of Agricultural Biotechnology, National Chung-Hsing University, Taichung, Taiwan.
Tsai
C C
CC
Tzen
J T
JT
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D002135:Calcium-Binding Proteins / Q000235:genetics*; D002352:Carrier Proteins / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010938:Plant Oils / Q000737:chemistry*; D010940:Plant Proteins; D010945:Plants, Edible / Q000737:chemistry* / Q000235:genetics*; D012313:RNA / Q000032:analysis; D029701:Saccharomyces cerevisiae Proteins; D012639:Seeds / Q000737:chemistry* / Q000235:genetics*
D048868:Adaptor Proteins, Signal Transducing; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D018076:DNA, Complementary; D010938:Plant Oils; D010940:Plant Proteins; C116430:SRO7 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C402836:caleosin protein, Sesamum indicum; D012313:RNA