Globule Structures Research Articles

Improving fat globule structure to narrow metabolite gap between human milk and infant formulae.

We hypothesized that improving the fat globule structure of infant formulae based on the milk fat globule membrane (MFGM) would regulate metabolites and metabolic pathways, making it more similar to the metabolic properties of human milk. Therefore, we prepared infant formulae with different fat globule structures, including two model infant formulae (F1: fat globules surrounded by MFGM; F2: fat globules surrounded by protein) and one commercial infant formulae containing MFGM, and compared their metabolic differences with those of human milk. The number of differential metabolites between each sample and human milk reached 60 (F1), 132 (F2) and 126 (IF1). Glycerophosphocholines were screened as potential biomarkers to distinguish infant formulae from human milk. Compared with F2 and IF1, the enrichment of amino acid metabolism and lipid metabolism pathways was not significant between F1 and human milk. These results emphasized that F1 had the highest similarity to human milk in metabolic properties.

Utilization of carboxyl group-grafted molybdenum disulfide for enhancing the performance of thin-film nanocomposite nanofiltration membranes

The incompatibility between inorganic nanoparticles and polyamide matrix in thin film nanocomposite (TFN) nanofiltration membranes tends to generate defects in the selective layer and further sacrifice the membrane separation performance, which can be promisingly mitigated by modifying nanoparticles with hydrophilic functional groups. In this study, carboxyl group-grafted molybdenum disulfide (COOH-MoS2) nanoparticles were prepared via organic functionalization grafting reaction and then incorporated into membrane polyamide layers. The resultant TFN membranes exhibited a rougher surface with unique globule structures, which greatly contributed to the enhancement of water permeance. The optimal membrane obtained at the COOH-MoS2 depositing density of 50.0 μg/cm2 exhibited a high Na2SO4 rejection of 98.5 %, a low NaCl rejection of 23.8 %, and an excellent water permeance of 27.7 LMH/bar, which was 2.8 and 1.5 times higher than that of the control membrane (9.6 LMH/bar) and the original MoS2 modified TFN membrane (18.4 LMH/bar), respectively. The more superior membrane performance enabled by COOH-MoS2 was mainly due to the enhanced hydrophilicity of the nanoparticles, which effectively inhibited the piperazine monomer diffusion and resulted a thinner and properly looser selective layer. This work provides an effective strategy to improve the performance of TFN nanofiltration membranes via graft of proper functional groups on nanomaterials.

High Energy Channeling and Malleable Transition States: Molecular Dynamics Simulations and Free Energy Landscapes for the Thermal Unfolding of Protein U1A and 13 Mutants.

The spliceosome protein U1A is a prototype case of the RNA recognition motif (RRM) ubiquitous in biological systems. The in vitro kinetics of the chemical denaturation of U1A indicate that the unfolding of U1A is a two-state process but takes place via high energy channeling and a malleable transition state, an interesting variation of typical two-state behavior. Molecular dynamics (MD) simulations have been applied extensively to the study of two-state unfolding and folding of proteins and provide an opportunity to obtain a theoretical account of the experimental results and a molecular model for the transition state ensemble. We describe herein all-atom MD studies including explicit solvent of up to 100 ns on the thermal unfolding (UF) of U1A and 13 mutants. Multiple MD UF trajectories are carried out to ensure accuracy and reproducibility. A vector representation of the MD unfolding process in RMSD space is obtained and used to calculate a free energy landscape for U1A unfolding. A corresponding MD simulation and free energy landscape for the protein CI2, well known to follow a simple two state folding/unfolding model, is provided as a control. The results indicate that the unfolding pathway on the MD calculated free energy landscape of U1A shows a markedly extended transition state compared with that of CI2. The MD results support the interpretation of the observed chevron plots for U1A in terms of a high energy, channel-like transition state. Analysis of the MDUF structures shows that the transition state ensemble involves microstates with most of the RRM secondary structure intact but expanded by ~14% with respect to the radius of gyration. Comparison with results on a prototype system indicates that the transition state involves an ensemble of molten globule structures and extends over the region of ~1–35 ns in the trajectories. Additional MDUF simulations were carried out for 13 U1A mutants, and the calculated φ-values show close accord with observed results and serve to validate our methodology.

Exposure of Von Willebrand Factor Cleavage Site in A1A2A3-Fragment under Extreme Hydrodynamic Shear.

Von Willebrand Factor (vWf) is a giant multimeric extracellular blood plasma involved in hemostasis. In this work we present multi-scale simulations of its three-domains fragment A1A2A3. These three domains are essential for the functional regulation of vWf. Namely the A2 domain hosts the site where the protease ADAMTS13 cleavages the multimeric vWf allowing for its length control that prevents thrombotic conditions. The exposure of the cleavage site follows the elongation/unfolding of the domain that is caused by an increased shear stress in blood. By deploying Lattice Boltzmann molecular dynamics simulations based on the OPEP coarse-grained model for proteins, we investigated at molecular level the unfolding of the A2 domain under the action of a perturbing shear flow. We described the structural steps of this unfolding that mainly concerns the -strand structures of the domain, and we compared the process occurring under shear with that produced by the action of a directional pulling force, a typical condition of single molecule experiments. We observe, that under the action of shear flow, the competition among the elongational and rotational components of the fluid field leads to a complex behaviour of the domain, where elongated structures can be followed by partially collapsed melted globule structures with a very different degree of exposure of the cleavage site. Our simulations pose the base for the development of a multi-scale in-silico description of vWf dynamics and functionality in physiological conditions, including high resolution details for molecular relevant events, e.g., the binding to platelets and collagen during coagulation or thrombosis.

Comparative analysis of interfacial composition and structure of fat globules in human milk and infant formulas

In this study, we analyzed and compared the fat globule interfacial compositions and structures in human milk and three types of infant formulas (IF1: supplied with MFGM, IF2: without MFGM and soy lecithin, IF3: supplied with soy lecithin). The results suggested that the interfacial protein composition of IF1 was comparatively closer to human milk, but still lacked certain bioactive MFGM proteins including XO, ADPH and PAS6/7. Considering the interfacial phospholipid, we observed 23, 31, and 29 phospholipid species that could be used to distinguish human milk and infant formulas (IF1, IF2 and IF3). We also found that phosphatidylinositol (PI) and phosphatidylserine (PS), which can absorb lipase, phosphatidylethanolamine (PE), and phosphatidylcholine (PC) and connect functional fatty acids, were lacking in infant formulas. Moreover, the infant formulas had a smaller average particle size of 0.38 μm and a thicker interfacial layer that interacted with the casein micelles. Even if IF1 included MFGM, fat globules structures like those found in human milk did not form, and the majority of the MFGM was still in the aqueous phase in free form. Overall, the comprehensive analysis in this study could be used to simulate or mimic human milk lipids at the supramolecular level.

A Comparative Analysis of the Structure and Biological Properties of Films and Microfibrous Scaffolds Based on Silk Fibroin.

A comparative analysis of the structure and biological properties of silk fibroin constructions was performed. Three groups of constructions were obtained: films obtained by casting an aqueous solution of silk fibroin and electrospun microfibrous scaffolds based on silk fibroin, with the addition of 30% gelatin per total protein weight. The internal structures of the films and single fibers of the microfibrous scaffolds consisted of densely packed globule structures; the surface area to volume ratios and volume porosities of the microfibrous scaffolds were calculated. All constructions were non-toxic for cells and provide high levels of adhesion and proliferation. The high regenerative potential of the constructions was demonstrated in a rat full-thickness skin wound healing model. The constructions accelerated healing by an average of 15 days and can be considered to be promising constructions for various tasks of tissue engineering and regenerative medicine.

Effects of microfiltration combined with ultrasonication on shelf life and bioactive protein of skim milk

To extend the shelf life and retain bioactive proteins in milk, this study utilized microfiltration (MF) combined with ultrasonication to treat skim milk and investigated its efficiency in removing bacteria and retaining bioactive proteins compared with HTST pasteurization and microfiltration alone. Results showed that microfiltration combined with ultrasonication at 1296 J/mL could completely remove the bacteria in skim milk. Ultrasonication further extended the shelf life (4 °C) of microfiltered skim milk, which could reach at least 40 days when MF was combined with ˃1296 J/mL ultrasonication. In addition, ELISA showed that HTST pasteurization significantly decreased the levels of IgG by ~30%, IgA by ~ 50%, IgM by ~60%, and lactoferrin by ~40%, whereas the activity of the enzymes lactoperoxidase and xanthine oxidase were also decreased by ~ 20%. Compared with HTST, MF alone or combined with ultrasonication retained these bioactive proteins to a larger degree. On the other hand, proteomics indicated both damage to casein micelle and fat globule structures in milk when ultrasonication at >1296 J/mL was applied, as shown by increases in caseins and milk fat globular proteins. Simultaneously, this ultrasound intensity also decreased levels of bioactive proteins, such as complement factors. Taken together, this study provided new insights that may help to implement this novel combination of non-thermal technologies for the dairy industry aimed at improving milk quality and functionality.

Dermoscopic clues of palmoplantar hyperkeratotic eczema and palmoplantar psoriasis: A prospective, comparative study of 90 patients.

Diagnosis can be difficult in isolated palmar and plantar lesions in patients with psoriasis and eczema. The purpose of our study is to compare the dermoscopic findings in patients with palmoplantar psoriasis and palmoplantar hyperkeratotic eczema. This prospective, comparative study included 90 patients histopathologically diagnosed with eczema or psoriasis (35 psoriasis and 55 eczema). The age range was 18-75years. The most common vessel type was dot vessel in psoriasis. Red globular ring vessels were seen in five patients with psoriasis, but not in any with eczema (P=0.007). The most common vascular distribution pattern was regular in psoriasis (40%). Patchy vascular pattern was significant in eczema. The most common background color was light red in psoriasis (48.6%) (P<0.001). Brownish-orange globules were observed in 25.7% of patients with eczema and 5.7% in patients with psoriasis (P=0.02). There is only one study in the published work about dermoscopy of palmoplantar psoriasis and eczema. In our study, yellow crusts, patchy scale distribution, patchy vascular pattern, yellow scale color, dull red background color and brownish-orange globules were significant in patients with palmoplantar eczema. On the other hand, patients with psoriasis had light red background color, regular vascular distribution pattern and white scale color. We observed globule structures with a pale center and dark peripheral rim only in patients with eczema, which was not identified in previous studies. This globule structure may be a new finding in eczema.

Using the USAXS technique to reveal the fat globule and casein micelle structures of bovine dairy products.

Cows' milk is a commodity used worldwide to make many dairy products. We have used the ultra small angle X-ray scattering (USAXS) technique to reveal the fat globule and casein micelle structures of some dairy products. USAXS covers the q-range 5×10-4Å-1<q<10-1Å-1, thereby allowing the study of micron-scale structures present in those dairy products. We measured the USAXS intensity, Iq, as a function of the scattering vector magnitude, q, for samples of skim milk, non-homogenized whole milk, homogenized whole milk, half and half and heavy cream, at two temperatures, 7°C and 45°C. The data collected from the scattering experiments were fitted using the Unified fit model run under the IRENA software from the Advanced Photon Source, Argonne National Laboratory (Illinois, USA). The fittings were carried out when the data were plotted as log[I(q)] vs log[q]. We observed a combination of linear regions (LRs) and knees. Two parameters of interest were obtained from the fittings, a radius of gyration, Rg, and a Porod exponent, P. Unified fit allowed us to fit up to four structural levels. One of the knees was centered at q ≈ 8×10-3Å-1 for all samples measured at 7°C, but vanished at 45°C. Two LRs were identified as being either due to casein micelles (CMs) or to fat globules (FGs). The porod exponent obtained from these LRs allowed us to describe the surface morphology of CMs and FGs. Two of the Rg values gave a rough estimate of the FGs and CMs sizes. FGs were identified for samples of homogenized whole milk, half and half and heavy cream in the q-region 2×10-4<q<8×10-4Å-1. We found that, in the absence of chymosin, or changes in pH, CaCl2 concentration or temperature changes, skim milk and non-homogenized whole milk displayed a Porod exponent that indicated a behavior characteristic of aggregation. Using computer simulations, we found that, seemingly, bovine CMs spontaneously formed approximately 1-dimensional aggregates possibly analogous to swollen randomly branched polymers.

Mechanical properties of C-S-H globules and interfaces by molecular dynamics simulation

At meso-scale, Calcium Silicate Hydrate (C-S-H) can be considered as randomly packed globules (about 4.2 nm), which forms the basic unit cell, with water molecules and voids. In this paper, the nanostructures for the globules are developed based on some plausible atomic structures of C-S-H. The mechanical properties for the C-S-H globules are determined through molecular dynamics simulation. Interfaces between the C-S-H globules are also simulated with different amount of water molecules. Key material parameters, e.g., Young’s modulus, strength and fracture energy, are obtained. It has been found that longer mean chain length of silicate tends to increase the strength of C-S-H and change the fracture behavior from brittle to ductile failure, in the chain length direction. In the other direction, however, silicate chains do not play an important role while interlayer structure matters. Moreover, pores in the C-S-H nanostructures can considerably reduce the strength of the globule structures in the normal direction to silicate chain but the weakening effect becomes substantially less in silicate chain direction. Further, it has been found that for all types of the interfaces between C-S-H globules, the interface with no extra water molecules has the greatest tensile/shear strength. The mechanical properties obtained in this paper for C-S-H nanostructures and interfaces could be necessary inputs to the meso-scale modelling of C-S-H via either granular mechanics, i.e., DEM, or continuum mechanics, i.e., FEM.

Emerging β-Sheet Rich Conformations in Supercompact Huntingtin Exon-1 Mutant Structures.

There exists strong correlation between the extended polyglutamines (polyQ) within exon-1 of Huntingtin protein (Htt) and age onset of Huntington's disease (HD); however, the underlying molecular mechanism is still poorly understood. Here we apply extensive molecular dynamics simulations to study the folding of Htt-exon-1 across five different polyQ-lengths. We find an increase in secondary structure motifs at longer Q-lengths, including β-sheet content that seems to contribute to the formation of increasingly compact structures. More strikingly, these longer Q-lengths adopt supercompact structures as evidenced by a surprisingly small power-law scaling exponent (0.22) between the radius-of-gyration and Q-length that is substantially below expected values for compact globule structures (∼0.33) and unstructured proteins (∼0.50). Hydrogen bond analyses further revealed that the supercompact behavior of polyQ is mainly due to the "glue-like" behavior of glutamine's side chains with significantly more side chain-side chain H-bonds than regular proteins in the Protein Data Bank (PDB). The orientation of the glutamine side chains also tend to be "buried" inside, explaining why polyQ domains are insoluble on their own.

Collapse Transition of Branched Polymers in Dilute Solutions: Telechelic Star vs. H‐Polymer

SummarySelf‐assembly in polymeric material is facilitated by the presence of active hetero‐atoms either along the backbone or as pendant groups. Self‐assembly leads to a rich variety of morphological pattern influencing various properties of polymeric materials. Presence of branching along the backbone chain also imparts remarkable change in polymeric properties. We report dynamic Monte Carlo (DMC) simulation study on solution behavior of branched polymers. We take multi‐arm telechelic star polymers (TSP, one branch point with terminal active groups) and H‐shaped branched polymers (homopolymers with two branch points), elucidating the driving mechanisms of collapse transition and morphological development. The presence of terminal active groups in TSP drives the collapse transition producing segregated and compact globule structure, on deteriorating solvent quality. On the other hand, presence of two branch points in H‐shaped polymers incorporates conformational heterogeneity (viz., entropically originated) and produces segregated globule structures. The terminal functional groups in TSP form aggregate, resembles to “watermelons” (WM) and double watermelon (DWM) depending on the number of arms and mode of cooling. In H‐polymers, conformational heterogeneity leads to the formation of segregated backbone and branch units (resembles to “Sandwich” or “Janus” morphology) rather an evenly distributed structure consisting of all the units, accompanying with a larger size of branches compared to the backbone.

Cohesin-dependent globules and heterochromatin shape 3D genome architecture in S. pombe.

Eukaryotic genomes are folded into three-dimensional structures, such as self-associating topological domains, the borders of which are enriched in cohesin and CCCTC-binding factor (CTCF) required for long-range interactions1-7. How local chromatin interactions govern higher-order folding of chromatin fibers and the function of cohesin in this process remain poorly understood. Here we perform genome-wide chromatin conformation capture (Hi-C) analysis8 to explore the high-resolution organization of the Schizosaccharomyces pombe genome, which despite its small size exhibits fundamental features found in other eukaryotes9. Our analyses of wild type and mutant strains reveal key elements of chromosome architecture and genome organization. On chromosome arms, small regions of chromatin locally interact to form “globules”. This feature requires a function of cohesin distinct from its role in sister chromatid cohesion. Cohesin is enriched at globule boundaries and its loss causes disruption of local globule structures and global chromosome territories. By contrast, heterochromatin, which loads cohesin at specific sites including pericentromeric and subtelomeric domains9-11, is dispensable for globule formation but nevertheless affects genome organization. We show that heterochromatin mediates chromatin fiber compaction at centromeres and promotes prominent interarm interactions within centromere-proximal regions, providing structural constraints crucial for proper genome organization. Loss of heterochromatin relaxes constraints on chromosomes, causing an increase in intra- and inter-chromosomal interactions. Together, our analyses uncover fundamental genome folding principles that drive higher-order chromosome organization crucial for coordinating nuclear functions.

Development of a Stable Virus-Like Particle Vaccine Formulation against Chikungunya Virus and Investigation of the Effects of Polyanions

Chikungunya virus (CHIKV) is an alphavirus that infects millions of people every year, especially in the developing world. The selective expression of recombinant CHIKV capsid and envelope proteins results in the formation of self-assembled virus-like particles (VLPs) that have been shown to protect nonhuman primates against infection from multiple strains of CHIKV. This study describes the characterization, excipient screening, and optimization of CHIKV VLP solution conditions toward the development of a stable parenteral formulation. The CHIKV VLPs were found to be poorly soluble at pH 6 and below. Circular dichroism, intrinsic fluorescence, and static and dynamic light scattering measurements were therefore performed at neutral pH, and results consistent with the formation of molten globule structures were observed at elevated temperatures. A library of generally recognized as safe excipients was screened for their ability to physically stabilize CHIKV VLPs using a high-throughput turbidity-based assay. Sugars, sugar alcohols, and polyanions were identified as potential stabilizers and the concentrations and combinations of select excipients were optimized. The effects of polyanions were further studied, and while all polyanions tested stabilized CHIKV VLPs against aggregation, the effects of polyanions on conformational stability varied.

Xylanase II from Trichoderma reesei QM 9414: conformational and catalytic stability to Chaotropes, Trifluoroethanol, and pH changes

Xylanase II, a key enzyme in the hydrolysis of xylan, was purified from cultures of Trichoderma reesei QM 9414 (anamorph of Hypocrea jecorina) grown on wheat straw as a carbon source. Xylanase treated with increasing guanidinium hydrochloride concentrations was denatured in a cooperative way regarding secondary and tertiary structures with midpoint transitions 5.6 ± 0.1 and 3.7 ± 0.1 M, respectively, whereas the enzymatic activity showed an intermediate state at 2-4 M denaturant. Treatment with urea showed that xylanase secondary structure was stabilized up to 4 M urea to be destabilized thereafter in a cooperative way with a transition midpoint Dm = 5.7 ± 0.2 M, but the ellipticity at 220 nm was greater than control in the presence of urea up to 6 M. Tertiary structure in the presence of urea showed also intermediate states with partial cooperative transitions with a midpoint: Dm = 2.7 ± 0.04 and 6.7 ± 0.3 M, respectively, whereas the enzymatic activity was enhanced about 40% at 2 M and inhibited above 4 M urea. Assays with the fluorescent probe 4,4'-bis-1-phenylamine-8-naphftalene sulfonate (bis-ANS) proved that the intermediate states had the characteristics of molten globule structures. The change of free energy for xylanase in absence of denaturants obtained from the spectral centre of mass (SCM) data at 298 K is = − 17 kJmol⁻¹ . In the presence of increasing trifluoroethanol (TFE), the enzyme gained α-helix content and lose tertiary structure and catalytic activity. Changes in pH (2-9) had practically no effect on the secondary structure of the enzyme, whereas the SCM values indicated that tertiary structure is maintained above pH 4. Bis-ANS binds to xylanase at pH 2 and 2.5 and in the presence of 30-40% TFE (v/v) characterizing molten globule states in those environmental conditions.

PH Shifting Alters Solubility Characteristics and Thermal Stability of Soy Protein Isolate and Its Globulin Fractions in Different pH, Salt Concentration, and Temperature Conditions

Soy protein isolate (SPI), beta-conglycinin (7S), and glycinin (11S) were subjected to pH-shifting treatments, that is, unfolding at pH 1.5 or 12.0 followed by refolding at pH 7.0, to induce molten globule structures. Treated samples were analyzed for protein solubility, thermal stability, and aggregation in 0, 0.1, and 0.6 M NaCl solutions at pH 2.0-8.0. The pH(12) shifting resulted in drastic increases (up to 2.5-fold) in SPI solubility in the pH 6.0-7.0 range, especially at 0 M NaCl. The pH(1.5) shifting had a generally lesser effect on solubility. 11S exhibited a solubility pattern similar to that of SPI, but the solubility of 7S was unaffected by pH shifting except at 0.6 M NaCl. The pH shifting, notably at pH 12.0, produced soluble, disulfide-linked polymers from 11S and reduced (P < 0.05) its enthalpy but not its temperature of denaturation. Soy proteins structurally altered by pH shifting had a reduced sensitivity to thermal aggregation.

Spectroscopy and atomic force microscopy of biomass

Scanning probe microscopy has emerged as a powerful approach to a broader understanding of the molecular architecture of cell walls, which may shed light on the challenge of efficient cellulosic ethanol production. We have obtained preliminary images of both Populus and switchgrass samples using atomic force microscopy (AFM). The results show distinctive features that are shared by switchgrass and Populus. These features may be attributable to the lignocellulosic cell wall composition, as the collected images exhibit the characteristic macromolecular globule structures attributable to the lignocellulosic systems. Using both AFM and a single case of mode synthesizing atomic force microscopy (MSAFM) to characterize Populus, we obtained images that clearly show the cell wall structure. The results are of importance in providing a better understanding of the characteristic features of both mature cells as well as developing plant cells. In addition, we present spectroscopic investigation of the same samples.

Processing methods to control silk fibroin film biomaterial features

Control of silk structural and morphological features is reported for fibroin protein films via all aqueous processing, with and without polyethylene oxide (PEO). Silk films with thicknesses from 500 nm to 50 μm were generated with controllable surface morphologies by employing soft-lithography surface patterning or by adjusting PEO concentrations. FTIR analysis indicated that water-annealing, used to cure or set the films, resulted in increased β-sheet and α-helix content within the films. Steam sterilization provided an additional control point by increasing β-sheet content, while reducing random coil and turn structures, yet retaining film transparency and material integrity. Increased PEO concentration used during processing resulted in decreased sizes of surface globule structures, while simultaneously increasing uniformity of these features. The above results indicate that both surface and bulk morphologies and structures can be controlled by adjusting PEO concentration. The combined tool set for controlling silk film geometry and structure provides a foundation for further study of novel silk film biomaterial systems. These silk film biomaterials have potential applicability for a variety of optical and regenerative medicine applications due to their optical clarity, impressive mechanical properties, slow degradability, and biocompatibility.

The supramolecular structure of milk fat influences plasma triacylglycerols and fatty acid profile in the rat

The digestion fate of milk fat depending on its supramolecular structure for a given dairy product composition has rarely been studied. To highlight differences of lipid digestion, we measured (i) the plasma triacylglycerol and cholesterol concentrations and (ii) the total plasma fatty acid profile of fasted rats force-fed with different dairy preparations; the three creams and the unemulsified preparation had a similar composition with different and controlled fat suprastructures. All preparations, manufactured in the laboratory from a given milk batch, contained 205 +/- 3 g . kg(-1) fat that was either fed (i) unemulsified consecutively to the skim milk phase, or as a cream with the following fat globule structures: (ii) native milk fat globules of approximately 4 microm covered with the native milk fat globule membrane (MFGM), (iii) small native milk fat globules of approximately 2 microm selected from the latter by microfiltration and covered by the MFGM, or (iv) fine homogenized fat droplets of approximately 1 microm covered mainly with caseins. The plasma triacylglycerol appearance was delayed for the creams compared with the rapid onset for the unemulsified preparation. At 90 and 180 min after feeding, the plasma triacylglycerol enrichment was significantly lower for the homogenized cream than for the unemulsified preparation. At 120 min after feeding, triacylglycerol enrichment was significantly lower for each cream than for the unemulsified preparation. At 180 min after feeding, the plasma relative enrichment in C12, C14, C15, C16 and C18:1 n-9 fatty acids was significantly lower for the homogenized cream than for unemulsified fat and regular cream. Global lipid digestion based on plasma triacylglycerol enrichment and relative enrichments in some fatty acids was decreased with small homogenized milk fat droplets compared to unemulsified milk fat. These data show that dairy products with the same composition but varying in fat supramolecular structure result in different kinetics of lipid digestion, which could be of health concern.

β-Lactoglobulin Molten Globule Induced by High Pressure

Beta-lactoglobulin (beta-LG) was treated with high hydrostatic pressure (HHP) at 600 MPa and 50 degrees C for selected times as long as 64 min. The intrinsic tryptophan fluorescence of beta-LG indicated that HHP treatment conditions induced a conformational change. HHP treatment conditions also promote a 3-fold increase in the extrinsic fluorescence of 1-anilinonaphthalene-8-sulfonate and a 2.6-fold decrease for cis-paraneric acid, suggesting an increase in accessible aromatic hydrophobicity and a decrease in aliphatic hydrophobicity. Far-ultraviolet circular dichroism (CD) spectra reveal that the secondary structure of beta-LG converts from native beta-sheets to non-native alpha-helices following HHP treatment, whereas near-ultraviolet CD spectra reveal that the native tertiary structure of beta-LG essentially disappears. Urea titrations reveal that native beta-LG unfolds cooperatively, but the pressure-treated molecule unfolds noncooperatively. The noncooperative state is stable for 3 months at 5 degrees C. The nonaccessible free thiol group of cysteine121 in native beta-LG became reactive to Ellman's reagent after adequate HHP treatment. Gel electrophoresis with and without beta-mercaptoethanol provided evidence that the exposed thiol group was lost concomitant with the formation of S-S-linked beta-LG dimers. Overall, these results suggest that HHP treatments induce beta-LG into hydrophobic molten globule structures that remain stable for at least 3 months.