Static Light Scattering Research Articles

Second Virial Coefficient As Determined from Protein Phase Behavior.

We quantitatively link the macroscopic phase behavior of protein solutions to protein-protein interactions based on a coarse-grained colloidal approach. We exploit the extended law of corresponding states and apply the Derjaguin-Landau-Verwey-Overbeek (DLVO) theory in order to infer the second virial coefficient b2, an integral measure of the interaction potential, from the phase behavior, namely, cloud-point temperature (CPT) measurements under conditions favoring protein crystallization. This determination of b2 yields values that quantitatively agree with the results of static light scattering (SLS) experiments. The strength of the attractions is quantified in terms of an effective Hamaker constant, which accounts for van der Waals attractions as well as non-DLVO forces, such as hydration and hydrophobic interactions. Our approach based on simple lab experiments to determine the CPT in combination with the DLVO theory is expected to facilitate further biophysical research on protein-protein interactions in complex solution environments.

Temperature dependence of solution properties of anionic polyelectrolyte-cationic surfactant complexes in ethanol

Abstract Stoichiometric polyelectrolyte-surfactant complexes, PSCs, were prepared from poly(acrylate), PA - , and poly(styrenesulfonate), PSS - , anions and dodecyltrimethylammonium cations, DTA + , and their properties were investigated by density and viscosity measurements in ethanol solutions in dependence on concentration and temperature. The measured densities were used to calculate the apparent molar volumes of the repeat units of DTAPA and DTAPSS complexes. Apparent molar volumes at infinite dilution, so as the reduced viscosities, in ethanol solutions are only weakly dependent on temperature. Through features typical for polyelectrolytes, viscosity curves suggest some small dissociation of complexes in ethanol. Light scattering, LS, measurements performed in the angular range from 50 to 150° did not reveal so called slow relaxation mode, which is typical for strongly charged polyelectrolytes in aqueous solutions without or with only a low amount of added salt. Size distributions were therefore calculated from the measured correlation functions. They were multimodal at all angles and for both PSCs, identifying 3 populations: particles with average hydrodynamic radii (i) below 10 nm, (ii) in the range 20–30 nm, and (iii) in the range 200–300 nm. Analysis of angular dependency of LS intensity for the largest particle population enabled the determination of the radius of gyration and the shape parameter ρ of these species. Large sizes, ρ values (ρ = 1.6 and 1.9 for DTAPA and DTAPSS, respectively), and the Kratky plots of the scattering data suggest that these particles are associates of more than one PSC (reverse micelle). Dynamic vapor sorption experiments with thin films of the studied PSCs show that the DTAPSS complex absorbs at the most 35% of water per its weight, whereas the DTAPA one absorbs considerably more (around 75%). Different structure of complexes was taken as responsible for large differences in water uptake.

Influence of aqueous dispersions in place of organic solvents during the synthesis of shape memory polyurethanes on their structure and properties

In this work, the effect of synthesizing shape memory polyurethanes in aqueous dispersions instead of in organic solvents on the structure and properties of the obtained polymers was investigated. Shape memory polyurethanes based on polycaprolactone diol and isophorone diisocyanate were synthesized by two routes: (1) aqueous dispersion (PU/SMWATER) and (2) dissolution in THF (PU/SMTHF). The samples were analyzed by infrared spectroscopy (FTIR), X‐ray diffraction (XRD), static light scattering (SLS), atomic force microscopy (AFM), differential scanning calorimetry (DSC), dynamic mechanical analysis (DMA), and mechanical tests. The aqueous dispersion method led to the production of polyurethanes with a higher degree of phase separation and a higher degree of crystallinity. The morphology of the obtained polyurethanes demonstrated that PU/SMWATER displays a structure with better defined phase separated domains. The polyurethanes exhibited similar average molar masses, soft segment glass transitions (Tgs) and mechanical properties. The lower degrees of phase separation and crystallinity of the PU/SMTHF led to lower values for the shape memory properties (shape recovery ratio (Rr)). The observed changes in the structure of the polyurethanes due to the replacement of organic solvent by an aqueous dispersion during their syntheses confirmed the preparation of shape memory polyurethanes with enhanced shape memory properties. POLYM. ENG. SCI., 57:432–440, 2017. © 2016 Society of Plastics Engineers

New insights about flocculation process in sodium caseinate-stabilized emulsions

Flocculation process was studied in emulsions formulated with 10wt.% sunflower oil, 2, 5 or 7.5wt.% NaCas, and with or without addition of sucrose (0, 5, 10, 15, 20 or 30wt.%). Two different processing conditions were used to prepare emulsions: ultraturrax homogenization or further homogenization by ultrasound. Emulsions with droplets with diameters above (coarse) or below (fine) 1μm were obtained. Emulsions were analyzed for droplet size distribution by static light scattering (SLS), stability by Turbiscan, and structure by confocal laser scanning microscopy (CLSM) and small angle X-ray scattering (SAXS). SAXS data were fitted by a theoretical model that considered a system composed of poly dispersed spheres with repulsive interaction and presence of aggregates. Flocculation behavior was caused by the self-assembly properties of NaCas, but the process was more closely related to interfacial protein content than micelles concentration in the aqueous phase. The results indicated that casein aggregation was strongly affected by disaccharide addition, hydrophobic interaction of the emulsion droplets, and interactions among interfacial protein molecules. The structural changes detected in the protein micelles in different environments allowed understanding the macroscopic physical behavior observed in concentrated NaCas emulsions.

Characterizing Reversible Protein Association at Moderately High Concentration Via Composition-Gradient Static Light Scattering

Analysis of weakly self-associating macromolecules at concentrations beyond a few g/L is challenging on account of the confounding effect of thermodynamic nonideality on the association signal. When the reversible association comprises only 1 or 2 oligomeric species in equilibrium with the monomer, the nonideality may be accounted for in a relatively rigorous manner, but if more association states are involved, the analysis becomes quite complex. We show that under reasonable assumptions, the nonideality in a composition-gradient static light scattering measurement may be accounted for in a simple fashion. The correction is applied to determining the stoichiometry and binding affinity of a protein previously characterized via sedimentation equilibrium and dynamic light scattering. The results of the new analysis are remarkably self-consistent and in line with the expectations for the form of self-association predicted previously from analysis of the surface residuals, establishing composition-gradient multi-angle static light scattering with nonideality corrections as a critical technology for characterizing associative interactions in concentrated solutions.

Semi-automated microplate monitoring of protein polymerization and aggregation

Static light scattering (SLS) is a commonly used technique for monitoring dynamics of high molecular weight protein complexes such as protein oligomers or aggregates. However, traditional methods are limited to testing a single condition and typically require large amounts of protein and specialized equipment. We show that a standard microplate reader can be used to characterize the molecular dynamics of different types of protein complexes, with the multiple advantages of microscale experimental volumes, semi-automated protocols and highly parallel processing.

Xyloglucans from flaxseed kernel cell wall: Structural and conformational characterisation

The structure of ethanol precipitated fraction from 1M KOH extracted flaxseed kernel polysaccharides (KPI-EPF) was studied for better understanding the molecular structures of flaxseed kernel cell wall polysaccharides. Based on methylation/GC–MS, NMR spectroscopy, and MALDI-TOF-MS analysis, the dominate sugar residues of KPI-EPF fraction comprised of (1,4,6)-linked-β-d-glucopyranose (24.1mol%), terminal α-d-xylopyranose (16.2mol%), (1,2)-α-d-linked-xylopyranose (10.7mol%), (1,4)-β-d-linked-glucopyranose (10.7mol%), and terminal β-d-galactopyranose (8.5mol%). KPI-EPF was proposed as xyloglucans: [Display omitted] The substitution rate of the backbone is 69.3%; R1 could be T-α-d-Xylp-(1→, or none; R2 could be T-α-d-Xylp-(1→, T-β-d-Galp-(1→2)-α-d-Xylp-(1→, or T-α-l-Araf-(1→2)-α-d-Xylp-(1→; R3 could be T-α-d-Xylp-(1→, T-β-d-Galp-(1→2)-α-d-Xylp-(1→, T-α-l-Fucp-(1→2)-β-d-Galp-(1→2)-α-d-Xylp-(1→, or none. The Mw of KPI-EPF was calculated to be 1506kDa by static light scattering (SLS). The structure-sensitive parameter (ρ) of KPI-EPF was calculated as 1.44, which confirmed the highly branched structure of extracted xyloglucans. This new findings on flaxseed kernel xyloglucans will be helpful for understanding its fermentation properties and potential applications.

Characterization of Clusters and Unimers in Associating Solutions of Chitosan by Dynamic and Static Light Scattering

Association of two chitosans of different degrees of acetylation (DA) is studied in dilute water/acid/salt solutions by combined analysis of static and dynamic light scattering. The influence of sodium salt anions Cl− and CH3COO− on conformation of macromolecules and properties of clusters was studied. At DA = 15 mol%, the macromolecules exist either in rigid‐rod (Rg/Rh = 2.7) or in coil (Rg/Rh = 1.8) conformations in the solvents containing NaCl and CH3COONa, respectively. Chitosans with higher DA = 28% dissolved in the solvent containing CH3COONa are in the collapsed state (Rg/Rh < 0.8), predicted by Dobrynin (Macromolecules, 2004). The clusters are of spherical form (0.8 < Rg/Rh < 1.2). CH3COONa promotes higher fractal dimensionality of clusters (2.2–2.4) than NaCl (1.9). Higher critical aggregation concentration is observed in NaCl. The other hydrodynamic methods (capillary viscometry, velocity sedimentation, and Tsvetkov diffusometry) do not detect the clusters, probably, because of their disruption during flow. image

Prediction of Protein Aggregation in High Concentration Protein Solutions Utilizing Protein-Protein Interactions Determined by Low Volume Static Light Scattering

The development of highly concentrated protein formulations is more demanding than for conventional concentrations due to an elevated protein aggregation tendency. Predictive protein-protein interaction parameters, such as the second virial coefficient B22 or the interaction parameter kD, have already been used to predict aggregation tendency and optimize protein formulations. However, these parameters can only be determined in diluted solutions, up to 20 mg/mL. And their validity at high concentrations is currently controversially discussed. This work presents a μ-scale screening approach which has been adapted to early industrial project needs. The procedure is based on static light scattering to directly determine protein-protein interactions at concentrations up to 100 mg/mL. Three different therapeutic molecules were formulated, varying in pH, salt content, and addition of excipients (e.g., sugars, amino acids, polysorbates, or other macromolecules). Validity of the predicted aggregation tendency was confirmed by stability data of selected formulations. Based on the results obtained, the new prediction method is a promising screening tool for fast and easy formulation development of highly concentrated protein solutions, consuming only microliter of sample volumes.

Hollow lignin azo colloids encapsulated avermectin with high anti-photolysis and controlled release performance

Lignin-based azo polymer (AL-azo-H) was prepared from alkali lignin (AL) and then used to prepare uniform colloids via self-assembly. Due to the unique UV-blocking performance, AL-azo-H colloids were applied to encapsulate photosensitive pesticide avermectin (AVM). The self-assembled structure of AL-azo-H colloids and their controlled release and anti-photolysis performances for AVM were investigated by dynamic light scattering (DLS), static light scattering (SLS), transmission electron microscopy (TEM) and high performance liquid chromatography (HPLC). Results show that AL-azo-H colloids are hollow spheres. Hollow AL-azo-H colloidal spheres have remarkable AVM encapsulation efficiency as high as 61.49% (w/w). The cumulative release amount of AVM encapsulated by AL-azo-H (AL-azo-H@AVM) after 120h was 84% and the release was still going on. The decomposition rate of AL-azo-H@AVM was less than 41% even after 72h of UV irradiation. Hollow AL-azo-H colloidal spheres showed excellent UV-blocking and controlled release performance for AVM.

Site-Specific Hydrolysis Reaction C-Terminal of Methionine in Met-His during Metal-Catalyzed Oxidation of IgG-1.

The metal-catalyzed oxidation by [Fe(II)(EDTA)](2-)/H2O2 of IgG-1 leads to the site-specific hydrolysis of peptide bonds in the Fc region. The major hydrolytic cleavage occurs between Met428 and His429, consistent with a mechanism reported for the site-specific hydrolysis of parathyroid hormone (1-34) between Met8 and His9 (Mozziconacci, O.; et al. Mol. Pharmaceutics 2013, 10 (2), 739-755). In IgG-1, to a lesser extent, we also observe hydrolysis reactions between Met252 and Ile253. After 2 h of oxidation (at pH 5.8, 37 °C) approximately 5% of the protein is cleaved between Met428 and His429. For comparison, after 2 h of oxidation, the amount of tryptic peptides containing a Met sulfoxide residue represents less than 0.1% of the protein. The effect of this site-specific hydrolysis on the conformational stability and aggregation propensity of the antibody was also examined. No noticeable differences in structural integrity and conformational stability were observed between control and oxidized IgG-1 samples as measured by circular dichroism (CD), fluorescence spectroscopy, and static light scattering (SLS). Small amounts of soluble and insoluble aggregates (3-6%) were, however, observed in the oxidized samples by UV-visible absorbance spectroscopy and size exclusion chromatography (SEC). Over the course of metal-catalyzed oxidation, increasing amounts of fragments were also observed by SEC. An increase in the concentration of subvisible particles was detected by microflow imaging (MFI).

Structure of cellulose/direct dye complex regenerated from supercritical water

The regeneration of cellulose from supercritical water in the presence of direct dyes was studied by small- and wide-angle synchrotron X-ray scattering and cryo-transmission electron microscopy to understand the effects of dyes on the structure formation of cellulose. In addition, the interactions between cellulose and the direct dyes were characterized using molecular dynamics simulations. Peaks corresponding to cellulose II crystals were observed in the wide-angle X-ray diffraction pattern of cellulose regenerated from supercritical water without dyes, whereas these peaks were not observed in the diffraction patterns of samples with direct dye (Direct Red 28 or Direct Blue 1). This result indicated that the direct dyes prevented the crystallization of regenerated cellulose. The results of the molecular dynamics simulations indicated that the planes of glucose rings interacted with the aromatic moieties of the dyes and that the sulfonate groups of the dye molecules interacted with the hydroxyl groups of cellulose. In addition, the CH groups of the glucose rings and aromatic moieties of the dyes (e.g., naphthalene and biphenyl moieties) interacted weekly. When cellulose regenerates from solution, cellulose sheet structures formed via hydrophobic interactions appear as the initial structure. The direct dyes were found to affect the formation of this cellulose sheet structure because cellulose molecularly dissolved in supercritical water. In the Kratky plots for small-angle X-ray scattering, a peak was clearly observed for the cellulose and cellulose/DR28 samples in the region of smaller q (<0.5), indicating that the nanoscale assembly structures dispersed in these systems. Bundled sheet-like and twisted ribbon-like structures were observed in the supernatants of the cellulose and cellulose/DR28 samples. These dispersed structures were considered to be intermediates in the structural formation of cellulose.

Estimation from moments measurements for amyloid depolymerisation

Estimating reaction rates and size distributions of protein polymers is an important step for understanding the mechanisms of protein misfolding and aggregation, a key feature for amyloid diseases. This study aims at setting this framework problem when the experimental measurements consist in the time-dynamics of a moment of the population (i.e. for instance the total polymerised mass, as in Thioflavin T measurements, or the second moment measured by Static Light Scattering). We propose a general methodology, and we solve the problem theoretically and numerically in the case of a depolymerising system. We then apply our method to experimental data of depolymerising oligomers, and conclude that smaller aggregates of ovPrP protein should be more stable than larger ones. This has an important biological implication, since it is commonly admitted that small oligomers constitute the most cytotoxic species during prion misfolding process.

PH-Responsive Intra- and Inter-Molecularly Micelle Formation of Anionic Diblock Copolymer in Water

Poly(sodium2-(acrylamido)-2-methylpropanesulfonate)-block-poly(sodium11-(acrylamido)undecanoate) (PAMPS–PAaU) was synthesized via reversible addition-fragmentation chain transfer (RAFT)-controlled radical polymerization. The “living” polymerization of PAaU was evidenced by the fact that the molecular weight distribution was narrow (Mw/Mn = 1.23). The pH-induced association behavior of PAMPS–PAaU in 0.1 M NaCl aqueous solutions as a function of solution pH was investigated by 1H NMR spin-spin relaxation time, dynamic light scattering (DLS), static light scattering (SLS), and fluorescence probe techniques. These results indicated that PAMPS–PAaU formed polymer micelles in 0.1 M NaCl aqueous solutions at pH < 9. At pH = 8–9, the polymer formed the micelles intramolecularly due to hydrophobic self-association of the PAaU block within the single polymer chain. On the other hand, at pH < 8, micellization occurred intermolecularly to form polymer micelles comprising hydrophobic PAaU cores and hydrophilic PAMPS shells.

The Debye light scattering equation’s scaling relation reveals the purity of synthetic dendrimers

Spherical dendrimer structures cannot be structurally modeled using conventional polymer models of random coil or rod-like configurations during the calibration of the static light scattering (LS) detectors used to determine the molecular weight (M.W.) of a dendrimer or directly assess the purity of a synthetic compound. In this paper, we used the Debye equation-based scaling relation, which predicts that the static LS intensity per unit concentration is linearly proportional to the M.W. of a synthetic dendrimer in a dilute solution, as a tool to examine the purity of high-generational compounds and to monitor the progress of dendrimer preparations. Without using expensive equipment, such as nuclear magnetic resonance or mass spectrometry, this method only required an affordable flow injection set-up with an LS detector. Solutions of the purified dendrimers, including the poly(amidoamine) (PAMAM) dendrimer and its fourth to seventh generation pyridine derivatives with size range of 5–9 nm, were used to establish the scaling relation with high linearity. The use of artificially impure mixtures of six or seven generations revealed significant deviations from linearity. The raw synthesized products of the pyridine-modified PAMAM dendrimer, which included incompletely reacted dendrimers, were also examined to gauge the reaction progress. As a reaction toward a particular generational derivative of the PAMAM dendrimers proceeded over time, deviations from the linear scaling relation decreased. The difference between the polydispersity index of the incompletely converted products and that of the pure compounds was only about 0.01. The use of the Debye equation-based scaling relation, therefore, is much more useful than the polydispersity index for monitoring conversion processes toward an indicated functionality number in a given preparation.

Self-Association of Apo A-1 Studied with Dynamic and Static Light Scattering.

Static and dynamic light scattering were employed to determine simultaneously the average relative molecular mass, Mr, and the average hydrodynamic radius, Rh, of protein molecules. The new method was applied to the association-dissociation equilibrium of apolipoprotein A-1 (Apo A-1) and its thermal unfolding. As a control, lysozyme was measured as a nonassociating protein. Apo A-1 forms oligomers as a function of concentration and temperature, and the equilibrium can be described by a cooperative association model, consisting of a nucleation step and a growth step. At concentrations of 1 and 2.7 mg/mL, the Apo A-1 solution contained mainly monomers and octamers, with intermediates occurring at very low concentrations. Oligomer formation was maximal at 22 °C and was characterized by a temperature-dependent association constant. The cooperative association model allows the quantitative analysis of both the average relative molecular mass, Mr, and the average hydrodynamic radius, Rh, with the same set of model parameters which, in turn, are also applicable to analytical ultracentrifugation experiments. The light scattering experiments were reversible as long as the Apo A-1 solution was not heated above 60 °C.

Multimodal Dispersion of Nanoparticles: A Comprehensive Evaluation of Size Distribution with 9 Size Measurement Methods.

Evaluation of particle size distribution (PSD) of multimodal dispersion of nanoparticles is a difficult task due to inherent limitations of size measurement methods. The present work reports the evaluation of PSD of a dispersion of poly(isobutylcyanoacrylate) nanoparticles decorated with dextran known as multimodal and developed as nanomedecine. The nine methods used were classified as batch particle i.e. Static Light Scattering (SLS) and Dynamic Light Scattering (DLS), single particle i.e. Electron Microscopy (EM), Atomic Force Microscopy (AFM), Tunable Resistive Pulse Sensing (TRPS) and Nanoparticle Tracking Analysis (NTA) and separative particle i.e. Asymmetrical Flow Field-Flow Fractionation coupled with DLS (AsFlFFF) size measurement methods. The multimodal dispersion was identified using AFM, TRPS and NTA and results were consistent with those provided with the method based on a separation step prior to on-line size measurements. None of the light scattering batch methods could reveal the complexity of the PSD of the dispersion. Difference between PSD obtained from all size measurement methods tested suggested that study of the PSD of multimodal dispersion required to analyze samples by at least one of the single size particle measurement method or a method that uses a separation step prior PSD measurement.

Study on the forming condition and mechanism of the β conformation in poly (9,9-dioctylfluorene) solution

In this work, the forming condition and mechanism of the β conformation in poly (9,9-dioctylfluorene) (PFO) solution are studied. We find the forming condition of β conformation is strongly associated with the proportion of the poor solvent (ethanol) in mixed solution (toluene/ethanol). When the proportion of poor solvent is less than 40% (volume ratio), there is no β conformation in solution; but if the proportion of poor solvent increases to 40%, the β conformation starts to appear. Static and Dynamic Light Scattering research reveal that the molecular chains become collapsed; meanwhile the degree of aggregation is also enhanced when the ethanol proportion increases and proved by TEM. By changing the PFO concentration, we find the concentrations have great influence on the chains aggregation but little influence on the content of β conformation. So we infer the β conformation was formed not in solution but in aggregation, the forming reason was proved to be both intrachain interaction in single chain itself as well as interchain intraction with neighbors in aggregation, which were all enhanced by adding ethanol proportion. The study has a potential meaning to make optoelectronic films whose the condensed state structure was mostly decided by the precursor solution with high charge carrier mobility and devices efficiency by controlling the content of β conformation in PFO precursor solution.

Characterizing protein-protein Nonspecific Interactions by Static Light Scattering

Nonspecific interactions between biomacromolecules have long been recognized as playing important roles in the thermodynamics and kinetics of biochemical reactions in cellular environments. Our previous study found that nonspecific interactions with bovine serum albumin (BSA) reduced the affinity of maltose binding protein (MBP) for its ligand maltose [1]. To probe the nonspecific interactions further, here we turned to second virial coefficient (B22) measurements. B22 contains information about the nature and strength of intermolecular interactions. Previous measurements have largely focused on self-interactions. We recently developed a method based on static light scattering that probes both self- and mutual interactions in a binary mixturesm[2]. We are applying this method to MBP-BSA mixtures under a wide range of solvent conditions (e.g., pH and salt concentration). We are also using maltose binding and mutations to induce conformational changes and modify the surface of MBP. At a neutral pH, our B22 results for the self- and mutual-interactions of MBP and BSA are all close to those predicted by pure steric repulsion, indicating that the net effects of soft interactions (i.e., electrostatic interaction and nonpolar attraction) for MBP-BMP, BSA-BSA, and MBP-BSA pairs are all nearly null. At pH 5, which is close to the isoelectric points of both proteins and hence charge-charge repulsion should be weakened, our results show moderately attractive soft interactions for the MBP-MBP and BSA-BSA pairs but strongly attractive soft interactions for the MBP-BSA pair. These measurements will provide valuable data for our computational efforts in parameterizing protein-protein interaction energy functions. Together, our experimental and computational studies will lead to quantitative understanding of protein-protein nonspecific interactions and how they influence biochemical reactions in cellular environments.[1] Wu D,et al.,J Phys Chem B, 2015, 119, 1891-8.[2] Miklos AC, et al., PLoS ONE 8(10): e74969.

Structural characterisation of outer membrane proteins from Borrelia burgdorferi sensu lato by small-angle X-ray scattering

Forming the interface between the bacterial cell and the host, the outer membrane of Borrelia is known to play a key role in pathogenicity. Although Borrelia burgdorferi sensu lato are considered to be Gram-negative, their outer membrane is unique, lacking liposaccharides and phosphatidylethanolamine. It contains a variety of glycolipids, surface exposed lipoproteins and a number of membrane-spanning β-barrels. BAPKO_0422, BB_0562 and BG_0408 are membrane proteins, theoretically predicted to form 8-stranded, membrane-spanning β-barrels. The aim of this work is to produce recombinant versions of these proteins, and determine molecular envelopes by small-angle X-ray scattering (SAXS). The β-barrel model can then be tested by comparing the experimental molecular envelope with the theoretical predictions. Three Borrelia proteins (BAPKO_0422, BB_0562 and BG_0408) were recombinantly expressed in the E. coli expression system using the pET-47 expression vector. The putative membrane proteins were purified by immobilised metal affinity chromatography (IMAC). The His tag of BAPKO_0422 was enzymatically removed to produce a native protein, and to allow for a visual comparison of the protein both with and without the His tag. SAXS data for each protein were collected and the overall shape was determined using ab initio methods. The pair-distance distribution function (P(r) function) of BAPKO_0422 with the His tag indicated a particle overlap potentially caused by the flexible 6-His tag at the N-terminus. Kratky plots of BAPKO_0422, BB_0562 and BG_0408 revealed the parabolic convergence for a folded particle. The low-resolution molecular envelopes of BAPKO_0422, BB_0562 and BG_0408 are consistent with the structure of an 8-stranded β-barrel. The filtered envelopes are in agreement with the shape and size of the E. coli homologue OmpX. The likely orientation of the protein within the outer membrane can be deduced by comparing molecular envelopes with and without the N-terminal His tag. The data suggest that BAPKO_0422, BB_0562 and BG_0408 are single-domain cylindrical-shaped molecules with no evidence of an internal pore. Several questions remain to be answered, such as the oligomeric state of BG_0408 and BAPKO_0422. The function of these 8-stranded β-barrels in the Borrelial outer membrane remains to be investigated.