Size Of Oil Bodies Research Articles

Study on the Stability Mechanism of Peanut OBs Extracted with the Aqueous Enzymatic Method.

In this study, the internal relationships among oil bodies (OBs), the protein-phospholipid interactions in aqueous phase, oil-water interface behavior, and the stability of reconstituted OBs were analyzed from the bulk phase, interface, and macro perspectives, and the stability mechanism of OBs was discussed. OB proteins and phospholipids were combined through hydrophobic and electrostatic interactions, resulting in the stretching of protein conformation. OB proteins and phospholipids act synergistically to increase interface pressure and the rate of increase in interface pressure with relatively stable elastic behavior, which is beneficial to the formation and stability of interfacial films. When OBs were reconstituted by an OB protein-phospholipid complex system, phospholipids bound to OB proteins through hydrophobic and electrostatic interactions. OB proteins and phospholipids uniformly covered the oil droplet surface of reconstituted OBs to form a stable interfacial film, which maintained the stability of OBs. The addition of phospholipids significantly reduced the particle size of OBs prepared by OB proteins in a dose-dependent manner, and particle size decreased with the increase in phospholipid content (p < 0.05). Phospholipids increased the net surface charge, enhanced electrostatic repulsion, and improved the physicochemical stability of reconstituted OBs. The stability mechanism elucidated in this study provides a theoretical basis for the demulsification of peanut OBs.

Characterization of oil body-associated proteins obtained from oil bodies with different sizes in oleaginous diatom Fistulifera solaris

Oil body-associated proteins from the oleaginous diatom Fistulifera solaris were identified by proteomic analysis of oil bodies of various sizes (small, middle, and large) by time-dependent culturing upon nutrient-starvation at 36, 96 and 168h. This diatom strain has the capability to accumulate neutral lipids and triacylglycerol. Liquid chromatography-tandem mass spectrometry analysis revealed 662 proteins in all oil body sizes. Among these, 132 proteins were predicted to be localized to the endoplasmic reticulum. Seventeen proteins that exhibited a positive correlation with gene expression and the oil body size were selected as novel candidates for oil body-associated proteins. Among the 17 protein candidates, two proteins encoded by fso:g8246 and fso:g10200 were confirmed to be localized on the surface of the oil body and endoplasmic reticulum. A protein encoded by fso:g2514, which is involved in sterol biosynthesis, was also identified. This protein was likely to localize to mitochondria; however, inhibitor assays suggested that it might play a role in lipid degradation. Our work provides new insights into the proteomics of microalgae and provides a valuable strategy for boosting lipid productivity in microalgae.

Characterization of Oil Body and Starch Granule Dynamics in Developing Seeds of Brassica napus.

Brassica napus is the most important oilseed crop in the world, and the lipid was stored in the oil body (OB) in the form of triacylglycerol. At present, most of studies on the relationship between oil body morphology and seed oil content in B. napus was focused on mature seeds. In the present study, the OBs in different developing seeds of B. napus with relatively high oil content (HOC) of about 50% and low oil content (LOC) of about 39% were analyzed. It was revealed that the size of OBs was first increased and then decreased in both materials. And in late seed developmental stages, the average OB size of rapeseed with HOC was higher than that of LOC, while it was reversed in the early seed developmental stages. No significant difference was observed on starch granule (SG) size in HOC and LOC rapeseed. Further results indicated that the expression of genes that involved in malonyl-CoA metabolism, fatty acid carbon chain extension, lipid metabolism, and starch synthesis in the rapeseed with HOC was higher than that of rapeseed with LOC. These results give some new insight for understanding the dynamics of OBs and SGs in embryos of B. napus.

Transcriptome and Regional Association Analyses Reveal the Effects of Oleosin Genes on the Accumulation of Oil Content in Brassica napus.

Rapeseed stores lipids in the form of oil bodies. Oil bodies in the seeds of higher plants are surrounded by oleosins. Adjusting oleosin protein levels can prevent the fusion of oil bodies and maintain oil body size during seed development. However, oil contents are affected by many factors, and studies on the complex molecular regulatory mechanisms underlying the variations in seed oil contents of B. napus are limited. In this study, a total of 53 BnOLEO (B. napus oleosin) genes were identified in the genome of B. napus through a genome-wide analysis. The promoter sequences of oleosin genes consisted of various light-, hormone-, and stress-related cis-acting elements, along with transcription factor (TF) binding sites, for 25 TF families in 53 BnOLEO genes. The differentially expressed oleosin genes between two high- and two low-oil-content accessions were explored. BnOLEO3-C09, BnOLEO4-A02, BnOLEO4-A09, BnOLEO2-C04, BnOLEO1-C01, and BnOLEO7-A03 showed higher expressions in the high-oil-content accessions than in low-oil-content accessions, at 25, 35, and 45 days after pollination (DAP) in two different environments. A regional association analysis of 50 re-sequenced rapeseed accessions was used to further analyze these six BnOLEO genes, and it revealed that the nucleotide variations in the BnOLEO1-C01 and BnOLEO7-A03 gene regions were related to the phenotypic variations in seed oil content. Moreover, a co-expression network analysis revealed that the BnOLEO genes were directly linked to lipid/fatty acid metabolism, TF, lipid transport, and carbohydrate genes, thus forming a molecular network involved in seed oil accumulation. These favorable haplotypes can be utilized in molecular marker-assisted selection in order to further improve seed oil contents in rapeseed.

Effect of extraction pH and post-extraction heat treatment on the composition and interfacial properties of peanut oil bodies

Oil bodies (OBs) are composed of internal triglyceride and external phospholipids with embedded surface proteins. The phospholipids and surface proteins not only play an important role in the stability of the natural OB droplets, but also endow emulsifying property of OBs, as they can participate in the emulsification process of a formula. In order to investigate the effects of extraction pH and post heat treatment on the interfacial properties of OBs, the interfacial composition and structure of peanut OBs extracted at different pHs, and heat treated at 30 ℃, 60 ℃, 90 ℃ after extracted at pH 7.5 were characterized, and their interfacial adsorption kinetics, interfacial isothermal curve, and viscoelasticity were studied using dilatational rheology and Langmuir-Blodgett (LB) film analyzer. The results showed that the content of peanut OBs surface proteins decreased with the increasing of extraction pH and heat treatment temperature. The particle size of peanut OBs extracted at pH 7.5 was the smallest and the surface hydrophobicity of these OBs heated at 30 ℃ was the highest, which resulted in their fastest increase of interfacial pressure and strongest viscoelasticity of the absorbed interfacial film at the oil-water interface. The peanut OBs extracted at different pH and heat-treatment demonstrated diffusion, adsorption and rupture/structural rearrangement at the air-water interface as time went on according to the interfacial compression isotherm curve, which are correlated with their interfacial dilatational rheology behaviors and emulsification ability. These findings provide a reference for the future utilization of peanut OBs as emulsifiers in the food and cosmetic industries.

Oleosin Expression Patterns and Size of Oil Bodies as a Factor in Determining Oil Content in Safflower (Carthamus Tinctorius L.) Genotypes

The climate crisis and the Ukraine war have shown the vulnerability of various crop commodities. One of those badly affected is cooking oil, leading to a shortage in several countries. This coupled with the need for healthier cooking oil, increases proportionally with the world population and has resulted in escalated cooking oil prices. Thus, continued evaluation of alternative oil crops that can do well in marginal lands becomes a vital practice to undertake. Safflower is one of the marginalized oil crops with high-quality oil containing essential fatty acids beneficial to human health. Screening safflower genotypes for oil content is critical for its breeding and adoption in non-native areas. Therefore, this study delineates the relationship between oleosin genes and oil bodies in regulating the oil content of safflower seeds. Oleosin genes and oil bodies from the seeds of five safflower genotypes were isolated and quantified using qPCR and fluorescence microscope respectively, and evaluated against the seed oil content. The results showed an inverse relationship where smaller oil bodies were displayed by genotypes with high oil content. A high relative expression of oleosin genes was observed in genotypes with high oil content (Kenya-9819 and Gila). Of the eight Ctoleosin genes that were studied, it was observed that Ctoleosin genes (1, 4, 6, 7, and 8) were highly reliable in characterizing safflower genotypes based on the oil content. Kenya-9819 and Gila genotypes were found to have high oil potential and this was confirmed by a higher accumulation of the oleosin gene. A high correlation coefficient between oleosin, oil content, and oil body was also observed in this study. The findings suggest that selected oleosin genes and oil bodies are important traits to consider when characterizing oil seed crops for oil content.

Effects of pH and temperature on the stability of peanut oil bodies: New insights for embedding active ingredients

As a natural emulsion, peanut oil bodies (OBs) are often used to embed drugs and natural active ingredients. To improve the utilization rate of peanut OBs, it is thus necessary to study their stability under different environmental conditions (pH, temperature). There were differences in the content and composition of peanut oil body interface protein (OBIP) at different pH values. When the pH increased, exogenous proteins were eluted and the endogenous protein content increased. By analyzing the particle size distribution, the secondary structure of OBIPs, and the fatty acid composition of lipids in OBs, it can be concluded that their stability increased with the pH, and the special protein–phospholipid membrane protected the lipids from damage. Upon heating OBs with different pH at 30–90 °C for 30 min, the particle size and zeta potential of OB (pH 6.8) increased, and the OBIPs did not change, reducing the stability of the OB. The particle size distribution, zeta potential, and interfacial protein composition of OB (pH 8, 9, 10, 11) did not change significantly, and their stability remained unchanged. Therefore, when OB is used to embed active ingredients, OB should be purified and the heating temperature should not exceed 90 °C.

Improvement of soybean product flavor and quality as affected by extraction of soybean oil bodies based on a soymilk model system

ABSTRACT Oil bodies (OBs) are lipid-storing organelles of plant seeds; they provide energy for seed germination and seedling growth by oxidative degradation as substrate. Soybean OBs are assumed to be involved in the enzymatic oxidation reaction of volatile flavor formation of soymilk. In order to establish a soymilk model system with soybean OBs as the enzymatic oxidation substrate to study the formation mechanism of soymilk flavor, the effects of extraction times and extraction pH on the composition, particle size and volatile flavor of soybean OBs extracted from raw soymilk were studied. The results showed that most of the extrinsic proteins on the surface of the OBs were removed with increasing extraction times. When the extraction pH increased from 7.0 to 11.0, the relative content of neutral lipids in OBs increased by 6.22%, and the relative content of protein and phospholipids in OBs decreased by 3.23% and 2.88%, respectively. The particle sizes of the OBs gradually decreased from 424 nm to 334 nm. The concentrations of volatile flavor components also gradually decreased from 129.29 μg/L to 52.75 μg/L. The integrity and stability of the OBs were compromised when the extraction pH was ≥12.0. The pH 11.0-OBs were the appropriate substrate for the soymilk model system because of their high purity and stability, better emulsion dispersion stability, and weaker flavor. These results are of great significance for exploring the formation mechanism of soymilk flavor in soymilk model systems to further improve the flavor of soymilk.

Association between Productivity, Fatty Acid Profiles, Oil Bodies’ Ultrastructure and Molecular Markers in Peanut (Arachis hypogaea L.) Cultivars

The commercial Nigerian peanut cultivars Ex-Dakar, Samnut-22 and Samnut-26, the Egyptian cultivar Ismailia-1 and the American cultivar NC-7 were genetically differentiated and the cultivars’ effects on growth and yield components, total lipid contents, including saturated and unsaturated fatty acids, and oil body size and number were investigated. Experiments were carried out at Cairo University Research Station, Giza, Egypt, during the summer seasons of 2016 and 2017. All cultivars had high contents of total unsaturated fatty acids and low contents of saturated fatty acids. Variations among tested cultivars in oil body size and number were documented and cultivars were ranked based on their oil bodies’ size as Ismailia-1 &gt; Samnut-26 &gt; NC-7 &gt; Ex-Dakar ≥ Samnut-22. Oil body size and number were inversely related. This variation can be explained by the different amounts of acidic amino acids and the protein structure on the surface of the oil bodies of these cultivars. The distributions of protein bodies and oil bodies seemed to be negatively correlated. Cultivar effect was significant, and variation was found in seed production and quality in addition to the size and number of oil bodies, oil productivity, and oil quality among cultivars. Seasonal effects were not evident due to the stability of the climatic conditions during the two seasons of the study.

The Development of Paeonia ostii Seeds and Oil Quality Formation

Paeonia ostii is a woody oil crop with potential value as an edible oil. With the aim of acquiring systematic knowledge of the development of P. ostii seeds, the oil content, biomass, and water content of the seeds were determined. Changes in the distribution of hydrogen protons in P. ostii seeds during follicle development were traced using magnetic resonance imaging (MRI). The formation of oil bodies in the endosperm and embryo was observed using transmission electron microscopy (TEM). Dynamic changes in oleic acid, linoleic acid, and α-linolenic acid contents were assessed by gas chromatography-mass spectrometry (GC-MS). The magnetic resonance images showed that, during early follicle development [45–85 days after flowering (DAF)], a greater quantity of liquid mucus was present within the seeds, and seeds in the same follicle developed at different rates. At 95 to 115 DAF, proton density was distributed evenly in all areas of the seed. A small dark area appeared in the center of the seed, and mucus in the follicles and water in the pericarp disappeared gradually. TEM observations showed that at 45 DAF, a few oil bodies were scattered at the cell periphery in the endosperm, and oil bodies were more numerous in the embryo. With the progression of seed development, the number and size of oil bodies in the embryo and endosperm continued to increase. The fresh and dry mass of P. ostii seeds increased from 45 to 105 DAF, then decreased after 105 DAF. The moisture content decreased, whereas the oil content increased and attained 33.1% at seed maturity. The three predominant unsaturated fatty acids accumulated simultaneously and showed stages of initial accumulation (45–65 DAF) and rapid accumulation (65–105 DAF). The results suggest that 65 to 105 DAF is a critical period for unsaturated fatty acid accumulation in P. ostii seeds.

Seed oil storage in three contrasted legume species: implications for oil improvement

Cellular comparisons on three legume species indicate oil body size is factor in mediating oil content. Key transcription factors and oleosins determining seed oil content were identified in somatic embryos of Medicago. Legume seeds provide a significant oil alternative to meat fat needs. Increasing demand for oil nutrition in the context of sustainable crop production has stimulated the exploration of legume seed oil storage regulation. This study investigated the cellular characteristics of seed oil storage using three legume species i.e. Medicago truncatula, Glycine max and Pongamia pinnata representing different oil/protein ratios, and then examined in vitro approaches for assessing strategies in enhancing seed oil storage. A greater range of oil body sizes was in higher oil/protein content species, with highest species having the largest oil bodies; and the smallest oil body size being relatively similar across species, suggesting that the arrangement of oil body size may be factor in mediating oil content. The expression of four key transcription factors i.e. LEC1, L1L, FUS3 and ABI3, and four oleosin genes in determining seed oil content was compared in vivo and in vitro using somatic embryos in Medicago, along with cellular evidence of oil bodies in somatic embryos, indicating that somatic embryos may be suitable models for rapid assessment of seed oil enhancement. This study revealed the cellular characteristics for legume seed oil storage with different nutritional compositions, and identified the associated molecular basis for boosting seed oil storage via regulating oil body size. In addition, somatic embryogenesis may be an effective system for examining oil production by modifying the expression of candidate genes prior to in vivo testing.

Improvement of Wax Oil Content of Embryonic Callus of Jojoba Using Gamma Radiation

Embryogenic callus was obtained from the leaf explants of jojoba (Simmondsia chinensis (Link) Schneider cultured on MS containing 0.5 mg/l NAA and 0.5 mg/l Kn. Growth of embryonic callus increased in 0.5 mg/l Kn + 6% sucrose. In order to improve oil content, the embryonic callus was exposed to different doses (5, 10 and 15 Kr) of gamma radiation. It was found that oil content of embryonic callus of jojoba increased 1.41% by exposing to 5 Kr gamma radiation. Also, size of oil bodies in embryonic callus irradiated with 5 Kr increased compared to control. Furthermore, production of the fatty acids was enhanced by gamma irradiation. The highest percentage of total fixed oil (1.41) was recorded in 5 Kr radiation. However, the highest content of total fatty acids (98.612) was obtained when embryonic callus were exposed to 10 Kr of gamma radiation.

Genome-wide identification and functional analysis of oleosin genes in Brassica napus L.

BackgroundRapeseed is the third largest oil seed crop in the world. The seeds of this plant store lipids in oil bodies, and oleosin is the most important structural protein in oil bodies. However, the function of oleosin in oil crops has received little attention.ResultsIn the present study, 48 oleosin sequences from the Brassica napus genome were identified and divided into four lineages (T, U, SH, SL). Synteny analysis revealed that most of the oleosin genes were conserved, and all of these genes experienced purifying selection during evolution. Three and four important oleosin genes from Arabidopsis and B. napus, respectively, were cloned and analyzed for function in Arabidopsis. Overexpression of these oleosin genes in Arabidopsis increased the seed oil content slightly, except for BnaOLE3. Further analysis revealed that the average oil body size of the transgenic seeds was slightly larger than that of the wild type (WT), except for BnaOLE1. The fatty acid profiles showed that the linoleic acid content (13.3% at most) increased and the peanut acid content (11% at most) decreased in the transgenic lines. In addition, the seed size and thousand-seed weight (TSW) also increased in the transgenic lines, which could lead to increased total lipid production.ConclusionWe identified oleosin genes in the B. napus genome, and overexpression of oleosin in Arabidopsis seeds increased the seed weight and linoleic acid content (13.3% at most).

Constitutive and Chloroplast Targeted Expression of Acetyl-CoA Carboxylase in Oleaginous Microalgae Elevates Fatty Acid Biosynthesis.

Photosynthetic microalgae are of burgeoning interest in the generation of commercial bioproducts. Microalgae accumulate high lipid content under adverse conditions, which in turn compromise their growth and hinder their commercial potential. Hence, it is necessary to engineer microalgae to mitigate elevated lipid accumulation and biomass. In this study, we identified acetyl-CoA carboxylase (ACCase) in oleaginous microalga Phaeodactylum tricornutum (PtACC2) and expressed constitutively in the chloroplast to demonstrate the potential of chloroplast engineering. Molecular characterization of transplastomic microalgae revealed that PtACC2 was integrated, transcribed and expressed successfully, and localized in the chloroplast. Enzymatic activity of ACCase was elevated by 3.3-fold, and the relative neutral lipid content increased substantially by 1.77-fold, and lipid content reached up to 40.8% of dry weight. Accordingly, the number and size of oil bodies markedly increased. Fatty acid profiling showed that the content of monounsaturated fatty acids increased, while polyunsaturated fatty acids decreased. This method provides a valuable genetic engineering toolbox for microalgal bioreactors with industrial significance.

Glucose-6-phosphate dehydrogenase as a target for highly efficient fatty acid biosynthesis in microalgae by enhancing NADPH supply

Oleaginous microalgae have great prospects in the fields of feed, nutrition, biofuel, etc. However, biomass and lipid productivity in microalgae remain a major economic and technological bottleneck. Here we present a novel regulatory target, glucose-6-phosphate dehydrogenase (G6PD) from the pentose phosphate pathway (PPP), in boosting microalgal lipid accumulation. G6PD, involved in the formation of NADPH demanded in fatty acid biosynthesis as reducing power, was characterized in oleaginous microalga Phaeodactylum tricornutum. In G6PD overexpressing microalgae, transcript abundance of G6PD increased by 4.4-fold, and G6PD enzyme activity increased by more than 3.1-fold with enhanced NADPH production. Consequently, the lipid content increased by 2.7-fold and reached up to 55.7% of dry weight, while cell growth was not apparently affected. The fatty acid composition exhibited significant changes, including a remarkable increase in monounsaturated fatty acids C16:1 and C18:1 concomitant with a decrease in polyunsaturated fatty acids C20:5 and C22:6. G6PD was localized to the chloroplast and its overexpression stimulated an increase in the number and size of oil bodies. Proteomic and metabolomic analyzes revealed that G6PD play a key role in regulating pentose phosphate pathway and subsequently upregulating NADPH consuming pathways such as fatty acid synthesis, thus eventually leading to lipid accumulation. Our findings show the critical role of G6PD in microalgal lipid accumulation by enhancing NADPH supply and demonstrate that G6PD is a promising target for metabolic engineering.

A pivotal role of vacuolar H(+)-ATPase in regulation of lipid production in Phaeodactylum tricornutum.

Microalgal lipids have been considered as a promising source for biodiesel production. Alkaline pH can induce neutral lipid accumulation in microalgae cells. However, whether and how proton pumps, especially vacuolar H+-ATPase (V-ATPase), function in these processes is not well known. In this study, we treated Phaeodactylum tricornutum with V-ATPase specific inhibitor bafilomycin A1 (BFA1) to determine its role in lipid production. Firstly, V-ATPase activity was increased in the latter phase of microalgae growth. BFA1 treatment decreased the cell density and lipid contents. Further analysis showed that BFA1 treatment reduced the number and size of oil bodies. GC-MS analysis showed that lipid components were not affected by BFA1 treatment. Intracellular pH was decreased and nitrogen depletion was delayed after BFA1 treatment. RNA-Seq analysis showed that expression of genes involved in calcium signaling, sulfur metabolism, cell cycle, glycolysis, pentose phosphate pathway, porphyrin, chlorophyll metabolism and lipid catabolic metabolism were upregulated, while expression of genes involved in ion transmembrane transport, ubiquitin mediated proteolysis, SNARE interactions in vesicular transport, fatty acid biosynthesis were downregulated under BFA1 treatment. Our findings provided insights into the molecular mechanisms underlying lipid accumulation and the key genes involved in lipid metabolism in Phaeodactylum tricornutum in response to BFA1.

Size and Charge Stability of Oil Bodies from Peanut

In order to offer scientific bases for the application of oil bodies from peanut in food, this research was undertaken to study the size and charge stability of oil bodies from five peanut varieties. It showed that the mean diameter of oil bodies fromyuhua9719andyuhua9830is obviously larger thanyuhua23,yuhua27, andyuhua9502in the peanut cell. Moreover, the analysis of diameter distribution of oil bodies also showed that the median diameter of oil bodies increased dramatically in the order ofyuhua9719&gt;yuhua9830&gt;yuhua23&gt;yuhua27&gt;yuhua9502after aqueous extraction. The charge stability of oil bodies from peanut was observed with zeta (ζ) potential measurements, which indicated that charge properties and the absolute value of oil bodies from five peanut varieties were significantly affected by pH and salt concentration, but the degree of influence is different. Of the five peanut varieties,yuhua27andyuhua9830possessed excellent charge stability (ζ-potential&gt;35 mV) in neutral microenvironment without salt concentration.

Behaviors of particle size and bound proteins of oil bodies in soymilk processing

In unheated soymilk, extrinsic proteins are bound to intact oil bodies coated by one monolayer of phospholipids and oil body intrinsic oleosins. In this study, effects of heating (70–100°C; 0–30min) on particle size and bound proteins of oil bodies were examined in suspension (oil bodies from unheated soymilk into deionized water) and soymilk. Mass ratio of extrinsic proteins/oleosins of oil bodies in unheated suspension and soymilk was respectively 1.1 and 2.5. By heating, extrinsic proteins released from oil bodies with different rates, and Z-average size of oil bodies increased in the beginning; afterwards, residual extrinsic proteins (extrinsic proteins/oleosins: 0.31 in suspension and 0.74 in soymilk; 100°C, 4min) and Z-average size were affected little. The amount of residual proteins at 80–100°C was negatively correlated with Z-average size of oil bodies in suspension (R2=0.996) and soymilk (R2=0.890).

Genome-Wide Analysis of Oleosin Gene Family in 22 Tree Species: An Accelerator for Metabolic Engineering of BioFuel Crops and Agrigenomics Industrial Applications?

Trees contribute to enormous plant oil reserves because many trees contain 50%-80% of oil (triacylglycerols, TAGs) in the fruits and kernels. TAGs accumulate in subcellular structures called oil bodies/droplets, in which TAGs are covered by low-molecular-mass hydrophobic proteins called oleosins (OLEs). The OLEs/TAGs ratio determines the size and shape of intracellular oil bodies. There is a lack of comprehensive sequence analysis and structural information of OLEs among diverse trees. The objectives of this study were to identify OLEs from 22 tree species (e.g., tung tree, tea-oil tree, castor bean), perform genome-wide analysis of OLEs, classify OLEs, identify conserved sequence motifs and amino acid residues, and predict secondary and three-dimensional structures in tree OLEs and OLE subfamilies. Data mining identified 65 OLEs with perfect conservation of the "proline knot" motif (PX5SPX3P) from 19 trees. These OLEs contained >40% hydrophobic amino acid residues. They displayed similar properties and amino acid composition. Genome-wide phylogenetic analysis and multiple sequence alignment demonstrated that these proteins could be classified into five OLE subfamilies. There were distinct patterns of sequence conservation among the OLE subfamilies and within individual tree species. Computational modeling indicated that OLEs were composed of at least three α-helixes connected with short coils without any β-strand and that they exhibited distinct 3D structures and ligand binding sites. These analyses provide fundamental information in the similarity and specificity of diverse OLE isoforms within the same subfamily and among the different species, which should facilitate studying the structure-function relationship and identify critical amino acid residues in OLEs for metabolic engineering of tree TAGs.

In vivo measurement of the size of oil bodies in plant seeds using a simple and robust pulsed field gradient NMR method

An easy to implement and convenient method to measure the mean size of oil bodies (OBs) in plant seeds is proposed using a pulsed field gradient nuclear magnetic resonance (PFGNMR) approach. PFGNMR is a well-known technique used to study either free or restricted diffusion of molecules. As triacylglycerols (TAG) are confined in OBs, analysis of their diffusion properties is a well-suited experimental approach to determine OB sizes. In fact, at long diffusion time, TAG mean squared displacement is limited by the size of the domain where these molecules are confined. In order to access the OB size distribution, strong intensities of magnetic field gradients are generally required. In this work we demonstrate for the first time that a standard liquid-phase NMR probe equipped with a weak-intensity gradient coil can be used to determine the mean size of OBs. Average sizes were measured for several seeds, and OB diameters obtained by PFGNMR were fully consistent with previously published values obtained by microscopy techniques. Moreover, this approach provided evidence of TAG transfer through the network of interconnected OBs, which is dependent on the ability of adjacent membranes to open diffusion routes between OBs. The main advantage of the NMR method is that it does not require any sample preparation and experiments are performed with whole seeds directly introduced in a standard NMR tube.