Guinier Analysis Research Articles

Lysozyme aggregation and unfolding in ionic liquid solvents: Insights from small angle X-ray scattering and high throughput screening

Understanding protein behaviour is crucial for developing functional solvent systems. Ionic liquids (ILs) are designer salts with versatile ion combinations, where some suppress unfavourable protein behaviour. This work utilizes small angle X-ray scattering (SAXS) to investigate the size and shape changes of model protein hen egg white lysozyme (HEWL) in 137 IL and salt solutions. Guinier, Kratky, and pair distance distribution analysis were used to evaluate the protein size, shape, and aggregation changes in these solvents. At low IL and salt concentration (1 mol%), HEWL remained monodispersed and globular. Most ILs increased HEWL size compared to buffer, while the nitrate and mesylate anions induced the most significant size increases. IL cation branching, hydroxyl groups, and longer alkyl chains counteracted this size increase. Common salts exhibited specific ion effects, while the IL effect varied with concentration due to complex ion-pairing. Protein aggregation and unfolding occurred at 10 mol% IL, altering the protein shape, especially for ILs with multiple alkyl chains on the cation, or with a mesylate/nitrate anion. This study highlights the usefulness of adopting a high-throughput SAXS strategy for understanding IL effects on protein behaviour and provides insights on controlling protein aggregation and unfolding with ILs.

Interrogating Encapsulated Protein Structure within Metal–Organic Frameworks at Elevated Temperature

Encapsulating biomacromolecules within metal-organic frameworks (MOFs) can confer thermostability to entrapped guests. It has been hypothesized that the confinement of guest molecules within a rigid MOF scaffold results in heightened stability of the guests, but no direct evidence of this mechanism has been shown. Here, we present a novel analytical method using small-angle X-ray scattering (SAXS) to solve the structure of bovine serum albumin (BSA) while encapsulated within two zeolitic imidazolate frameworks (ZIF-67 and ZIF-8). Our approach comprises subtracting the scaled SAXS spectrum of the ZIF from that of the biocomposite BSA@ZIF to determine the radius of gyration of encapsulated BSA through Guinier, Kratky, and pair distance distribution function analyses. While native BSA exposed to 70 °C became denatured, in situ SAXS analysis showed that encapsulated BSA retained its size and folded state at 70 °C when encapsulated within a ZIF scaffold, suggesting that entrapment within MOF cavities inhibited protein unfolding and thus denaturation. This method of SAXS analysis not only provides insight into biomolecular stabilization in MOFs but may also offer a new approach to study the structure of other conformationally labile molecules in rigid matrices.

Insights into Solution Structures of Photosynthetic Protein Complexes from Small-Angle Scattering Methods

High-resolution structures of photosynthetic pigment–protein complexes are often determined using crystallography or cryo-electron microscopy (cryo-EM), which are restricted to the use of protein crystals or to low temperatures, respectively. However, functional studies and biotechnological applications of photosystems necessitate the use of proteins isolated in aqueous solution, so that the relevance of high-resolution structures has to be independently verified. In this regard, small-angle neutron and X-ray scattering (SANS and SAXS, respectively) can serve as the missing link because of their capability to provide structural information for proteins in aqueous solution at physiological temperatures. In the present review, we discuss the principles and prototypical applications of SANS and SAXS using the photosynthetic pigment–protein complexes phycocyanin (PC) and Photosystem I (PSI) as model systems for a water-soluble and for a membrane protein, respectively. For example, the solution structure of PSI was studied using SAXS and SANS with contrast matching. A Guinier analysis reveals that PSI in solution is virtually free of aggregation and characterized by a radius of gyration of about 75 Å. The latter value is about 10% larger than expected from the crystal structure. This is corroborated by an ab initio structure reconstitution, which also shows a slight expansion of Photosystem I in buffer solution at room temperature. In part, this may be due to conformational states accessible by thermally activated protein dynamics in solution at physiological temperatures. The size of the detergent belt is derived by comparison with SANS measurements without detergent match, revealing a monolayer of detergent molecules under proper solubilization conditions.

Activated Carbon from Agricultural Wastes for Adsorption of Organic Pollutants.

Agricultural waste materials (strawberry seeds and pistachio shells) were used for preparation of activated carbons by two various methods. Chemical activation using acetic acid and physical activation with gaseous agents (carbon dioxide and water vapor) were chosen as mild and environmentally friendly methods. The effect of type of raw material, temperature, and activation agent on the porous structure characteristics of the materials was discussed applying various methods of analysis. The best obtained activated carbons were characterized by high values of specific surface area (555–685 m2/g). The Guinier analysis of small-angle X-ray scattering (SAXS) curves showed that a time of activation affects pore size. The samples activated using carbon dioxide were characterized mostly by the spherical morphology of pores. Adsorbents were utilized for removal of the model organic pollutants from the single- and multicomponent systems. The adsorption capacities for the 4-chloro-2-methyphenoxyacetic acid (MCPA) removal were equal to 1.43–1.56 mmol/g; however, for adsorbent from strawberry seeds it was much lower. Slight effect of crystal violet presence on the MCPA adsorption and inversely was noticed as a result of adsorption in different types of pores. For similar herbicides strong competition in capacity and adsorption rate was observed. For analysis of kinetic data various equations were used.

Solution Structure of the Detergent-Photosystem II Core Complex Investigated by Small-Angle Scattering Techniques.

Albeit achieving the X-ray diffraction structure of dimeric photosystem II core complexes (dPSIIcc) at the atomic resolution, the nature of the detergent belt surrounding dPSIIcc remains ambiguous. Therefore, the solution structure of the whole detergent-protein complex of dPSIIcc of Thermosynechococcus elongatus (T. elongatus) solubilized in n-dodecyl-ß-d-maltoside (ßDM) was investigated by a combination of small-angle X-ray scattering (SAXS) and small-angle neutron scattering (SANS) with contrast variation. First, the structure of dPSIIcc was studied separately in SANS experiments using a contrast of 5% D2O. Guinier analysis reveals that the dPSIIcc solution is virtually free of aggregation in the studied concentration range of 2-10 mg/mL dPSIIcc, and characterized by a radius of gyration of 62 Å. A structure reconstitution shows that dPSIIcc in buffer solution widely retains the crystal structure reported by X-ray free electron laser studies at room temperature with a slight expansion of the entire protein. Additional SANS experiments on dPSIIcc samples in a buffer solution containing 75% D2O provide information about the size and shape of the whole detergent-dPSIIcc. The maximum position of P(r) function increases to 68 Å, i.e., it is about 6 Å larger than that of dPSIIcc only, thus indicating the presence of an additional structure. Thus, it can be concluded that dPSIIcc is surrounded by a monomolecular belt of detergent molecules under appropriate solubilization conditions. The homogeneity of the ßDM-dPSIIcc solutions was also verified using dynamic light scattering. Complementary SAXS experiments indicate the presence of unbound detergent micelles by a separate peak consistent with a spherical shape possessing a radius of about 40 Å. The latter structure also contributes to the SANS data but rather broadens the SANS curve artificially. Without the simultaneous inspection of SANS and SAXS data, this effect may lead to an apparent underestimation of the size of the PS II-detergent complex. The formation of larger unbound detergent aggregates in solution prior to crystallization may have a significant effect on the crystal formation or quality of the ßDM-dPSIIcc.

Structure and dynamics analysis of a family 43 glycoside hydrolase α-L-arabinofuranosidase (PsGH43_12) from Pseudopedobacter saltans by computational modeling and small-angle X-ray scattering

The structure and molecular dynamics of α-L-arabinofuranosidase (PsGH43_12) of family 43 glycoside hydrolase, subfamily 12 from Pseudopedobacter saltans were studied. The modeled PsGH43_12 structure displayed 5-bladed β-propeller fold at N-terminal and β–sandwich fold at C terminal. Ramachandran plot showed 95.7% residues in favored and 3.3% in the generously allowed region and only 1% residues in the disallowed region. The secondary structure analysis of PsGH43_12 by circular dichroism revealed 2.7% α-helices, 30.33% β-strands and 66.97% random coils. Protein melting study of PsGH43_12 showed complete unfolding at 65°C and did not require any metal ion for its stability. Molecular docking analysis confirmed the involvement of active site residues Asp71, Asp180 and Glu247 in the catalysis, which was also confirmed by the site-directed mutagenesis of these residues. SAXS analysis displayed that PsGH43_12 is monomeric and a fully folded state in solution form. Guinier analysis gave the radius of gyration (Rg) 2.8 ± 0.09 nm. The maximum dimension and Rg of PsGH43_12 estimated from P(R) plot were 9.7 nm and 2.81 nm, respectively. The ab initio derived dummy model of PsGH43_12 displayed a bell-like shape. The ab initio derived dummy model superposed well with its comparative modeled structure except the N-terminal His6-tag region.

On the physico-chemical basis of self-nanosegregation giving magnetically-induced birefringence in dibutyl phosphate/bis(2-ethylhexyl) amine systems

Dibutyl phosphate (DBP)/bis(2-ethylhexyl) amine (BEEA) liquid mixtures can show optical birefringence when exposed to a magnetic field. It has been hypothesized that this is a consequence of self-segregation characterizing these systems giving formation of anisotropic local assemblies which are able to re-orient under magnetic field. In this work, an ab-initio Density Functional Theory computer modeling coupled with synchrotron X-Ray Scattering (XRS) experiments carried out at various BEEA molar ratio (X) and temperatures (10 ≤ T ≤ 60 °C) permitted to highlight the physical origin of this effect: a DBP-to-BEEA proton transfer with formation of an inherently anisotropic DBP-BEEA ion pair constituting the building block of highly structured ionic liquid nanodomains. At non-equimolar compositions (X ≠ 0.5) such ionic liquids domains are dispersed in the component in excess. At X > 0.5 the ionic liquids nanodomains are dispersed in excess of poorly structured BEEA, creating the ideal assumptions for the formation of a two-phase dispersed system, as confirmed by Guinier analysis in the small angle regime of the XRS spectra, explaining the system capability to become birefringent in reaction to external magnetic field.These information can be transferred to other amphiphiles-based mixtures for the piloted design of new magnetic field responsive materials with advanced applications.

Combined SAXS and computational approaches for structure determination and binding characteristics of Chimera (CtGH1-L1-CtGH5-F194A) generated by assembling β-glucosidase (CtGH1) and a mutant endoglucanase (CtGH5-F194A) from Clostridium thermocellum

Chimera (CtGH1-L1-CtGH5-F194A) developed by fusing β-glucosidase (CtGH1) at N-terminal and endoglucanase (CtGH5-F194A) at C-terminal was structurally characterized. Its secondary structure analysis by CD showed 38% α-helix, 9.3% β-sheets and 52.7% random coils corroborating with prediction. In-silico modeled structure of Chimera comprised two modules, CtGH1 and CtGH5-F194A displaying (α/β)8 fold. Ramachandran plot of Chimera showed 99.9% residues in allowed region. Binding interaction of Chimera with cello-oligosaccharides suggested active forms of CtGH1 and CtGH5-F194A and their involvement in catalysis. MD simulation of cellohexaose bound endoglucanase module of Chimera showed favourable flexibility in loops, LA with H-bond formation with Asn510 and in loop LC relocation of Tyr687 away from active site efficiently releasing the product after catalysis. Higher short range interaction energy of Chimera, −383 kJ/mol than the individual endoglucanase, 254 kJ/mol against cellohexaose suggested higher efficient catalysis by Chimera. β-Glucosidase module of Chimera showed fluctuations in outer loops suggesting conformational changes that might be contributing to improved hydrolysis. SAXS analysis of Chimera displayed monodispersed state. Guinier analysis of Chimera showed globular shape (Rg= 3.15 ± 0.10 nm). Kratky plot confirmed fully folded and flexible behaviour in solution. Gasbor modeled structure of Chimera displayed an elongated structure with two modules having shape similar to bean-bag contour.

Determining Rg of IDPs from SAXS Data.

There is a great interest within the research community to understand the structure-function relationship for intrinsically disordered proteins (IDPs); however, the heterogeneous distribution of conformations that IDPs can adopt limits the applicability of conventional structural biology methods. Here, scattering techniques, such as small-angle X-ray scattering, can contribute. In this chapter, we will describe how to make a model-free determination of the radius of gyration by using two different approaches, the Guinier analysis and the pair distance distribution function. The ATSAS package (Franke et al., J Appl Crystallogr 50:1212-1225, 2017) has been used for the evaluation, and throughout the chapter, different examples will be given to illustrate the discussed phenomena, as well as the pros and cons of using the different approaches.

A light-scattering study of highly refractive, irregularly shaped MoS2 particles

We present measurements of light scattering intensity from aerosolized, micron sized, irregularly shaped, molybdenum disulfide (MoS2) particles in order to study the effects of a refractive index, m = n + iκ, with large real and imaginary parts. Light scattering was measured over a range of angles from 0.32° to 157°. Calibration was achieved by scattering with micron sized, spherical silica particles. Light scattering for both particle types was compared to theoretical Mie scattering calculations using size distributions determined by an aerodynamic particle sizer. Effects of the intensity weighted size distribution are discussed. We find that scattering by these irregularly shaped, highly refractive particles is well described by Mie scattering. We also find that when the quantity κkR, where kR = 2πR/λ is the size parameter, is greater than one, there is no enhancement in the backscattering. Finally, we show that Guinier analysis of light scattering by highly refractive particles yields intensity weighted mean sizes of reasonable accuracy for any shape.

Small-angle X-ray scattering based structure, modeling and molecular dynamics analyses of a family 5 glycoside hydrolase first endo-mannanase named as RfGH5_7 from Ruminococcus flavefaciens

Endo-β-1,4-mannanase named as RfGH5_7 from Ruminococcus flavefaciens cloned, expressed and purified earlier was structurally characterized in present study. The RaptorX modeled structure of RfGH5_7 showed a (β/α)8 Triose-phosphate Isomerase (TIM) barrel fold. The Ramachandran plot assessment of RfGH5_7 showed that all amino acids fall in allowed region except one, Asn22 in the disallowed region. The superposition of RfGH5_7 modeled structure with its nearest homologues revealed that Glu154 acts as proton donor while Glu249 acts as nucleophile. Secondary structure of RfGH5_7 through Circular Dichroism (CD) analysis revealed 33.5% α-helices, 17% β-strands and 49.5% random coils. Molecular Dynamic (MD) simulation showed Root Mean Square Deviation (RMSD), 0.67 nm and radius of gyration (Rg) between 1.9 nm and 1.85 nm. The binding interaction of mannotetraose on the surface of RfGH5_7 structure displayed polar interactions with His219, Tyr221, Trp278, Ser279 and Gly282 residues. Small-angle X-ray scattering (SAXS) analysis displayed the intact and monodispersed nature of the enzyme RfGH5_7. The radius of gyration (Rg) by Guinier analysis for globular shape was found to be 2.29 ± 0.09 nm and for rod-shape it was 0.95 ± 0.02 nm. Kratky plot confirmed that RfGH5_7 structure is compact and folded in solution. The ab initio derived dummy model of RfGH5_7 displayed single domain structure of yellow humped fish like shape. The RfGH5_7 modeled structure was well fitted with ab initio derived model from SAXS data. Communicated by Ramaswamy H. Sarma

Small-angle scattering model analysis of cage-like uranyl peroxide nanoparticles

Uranyl peroxide nanoparticles (NPs) with spherical cage structure form in aqueous medium by the assembly of uranyl hexagonal bipyramids. Morphological evolution of uranyl peroxide NPs prepared with different LiOH/UO2 molar ratios is studied by small-angle X-ray scattering. Guinier analysis indicated that the overall size of the uranyl peroxide NPs increased with storage time. Their structure is further analyzed by model fitting to a spherical shell particle. Analysis of the particle size shows that U24 particles containing 24 uranium atoms are formed in dispersion with Li/U = 7.7, and U28 particles are generated in dispersion with Li/U = 3.1. With increasing storage time, the interactions between uranyl peroxide NPs changes from repulsion to attraction for samples with Li/U ratio 5.5 and 7.7, leading to their agglomeration and precipitation. Certain amount of large aggregates remained in the dispersions in ambient condition for over one year, as confirmed by dynamic light scattering. This study gives insight into the structural evolution and interactions of uranyl peroxide NPs.

Soot aggregate sizing in an extended premixed flame by high-resolution two-dimensional multi-angle light scattering (2D-MALS)

The spatial distribution of soot aggregate size and morphology within a premixed flat flame (McKenna-type burner and ethyne–air mixture at an equivalence ratio of Φ = 2.7) is characterized by two-dimensional multi-angle light scattering (2D-MALS). A profound investigation of such an extended, radially symmetrical sooting flame with 2D-MALS requires a sophisticated camera calibration to correct for non-linear image scaling and a careful evaluation of the scattering data. Sharp scattering images were acquired in the angular range from 20° to 155° using a rotatable camera system and an automated Scheimpflug adapter. To correct for non-linear variations in horizontal and vertical image magnification occurring at scattering angles differing from perpendicular view, a polynomial-based image transformation algorithm was developed to convert all scattering images into a common coordinate system. Effective radii of gyration and fractal dimensions of soot aggregates were then derived from scattering data by two different approaches. Due to limited amount of angular positions, the classical method based on Guinier and power law analysis shows limitations, as it yields discontinuous results, predominantly in axial direction of the burner. Bayesian analysis was then used for a data fit of the complete structure factor conducting a least square minimization leading to more consistent results. The use of prior knowledge in the Bayesian evaluation allows for improved data fitting and reduced uncertainties in radius of gyration and fractal dimension even for small aggregate sizes.

Effect of Weakly Interacting Cosolutes on Lysozyme Conformations.

Exposure of a protein to cosolutes, like denaturants, changes its folding equilibrium. To determine the ensemble of protein conformations at equilibrium, in the presence of weakly interacting cosolutes, we present a two-stage analysis of solution X-ray scattering data. In the first stage, Guinier analysis and Kratky plot revealed information about the compactness and flexibility of the protein. In the second stage, elastic network contact model and coarse-grained normal mode analysis were used to generate an ensemble of conformations. The scattering curves of the conformations were computed and fitted to the measured scattering curves to get insights into the dominating folding states at equilibrium. Urea and guanidine hydrochloride (GuHCl) behaved as preferentially included weakly interacting cosolutes and induced denaturation of hen egg-white lysozyme, which served as our test case. The computed models adequately fit the data and gave ensembles of conformations that were consistent with our measurements. The analysis suggests that in the presence of urea, lysozyme retained its compactness and assumed molten globule characteristics, whereas in the presence of GuHCl lysozyme adopted random coiled conformations. Interestingly, no equilibrium intermediate states were observed in both urea and GuHCl.

In situ observation of synthesized nanoparticles in ultra-dilute aerosols via X-ray scattering

In-air epitaxy of nanostructures (Aerotaxy) has recently emerged as a viable route for fast, large-scale production. In this study, we use small-angle X-ray scattering to perform direct in-flight characterizations of the first step of this process, i.e., the engineered formation of Au and Pt aerosol nanoparticles by spark generation in a flow of N2 gas. This represents a particular challenge for characterization because the particle density can be extremely low in controlled production. The particles produced are examined during production at operational pressures close to atmospheric conditions and exhibit a lognormal size distribution ranging from 5–100 nm. The Au and Pt particle production and detection are compared. We observe and characterize the nanoparticles at different stages of synthesis and extract the corresponding dominant physical properties, including the average particle diameter and sphericity, as influenced by particle sintering and the presence of aggregates. We observe highly sorted and sintered spherical Au nanoparticles at ultra-dilute concentrations (< 5 × 105 particles/cm3) corresponding to a volume fraction below 3 × 10–10, which is orders of magnitude below that of previously measured aerosols. We independently confirm an average particle radius of 25 nm via Guinier and Kratky plot analysis. Our study indicates that with high-intensity synchrotron beams and careful consideration of background removal, size and shape information can be obtained for extremely low particle concentrations with industrially relevant narrow size distributions.

The Gelsolin Pathogenic D187N Mutant Exhibits Altered Conformational Stability and Forms Amyloidogenic Oligomers.

Gelsolin is an actin-severing protein that attains an open functional conformation in the presence of Ca2+ or low pH. Mutations (D187N/Y) in the second domain of gelsolin trigger the proteolytic pathway producing amyloidogenic fragments that form the pathological hallmark of gelsolin amyloidosis and lattice corneal dystrophy type 2 (LCD2). Here, we show that the D187N mutant gelsolin in a Ca2+ depleted, low pH-activated, open conformation could assemble into amyloidogenic oligomers without necessarily undergoing the specific proteolytic step. Although both wild-type (WT) and mutant proteins exhibit closely overlapping globular shapes at physiological conditions, the latter exhibits subjugated actin depolymerization, loss of thermodynamic stability, and folding cooperativity. Mutant gelsolin displayed aberrant conformational unwinding and formed structural conformers with high associative properties at low pH conditions. A SAXS intensity profile and Guinier analysis of these conformers showed the formation of unusual, higher order aggregates. Extended incubation at low pH resulted in the formation of thioflavin T and Congo red positive, β-sheet rich aggregates with a fibrillar, amyloid-like morphology visible under electron and atomic force microscopy. Mass spectrometric analysis of disaggregated end-stage fibrils displayed peptide fragments encompassing the entire protein sequence, indicating the involvement of full length mutant gelsolin in fibril formation. Atomistic and REMD simulations indicated a larger increase in solvent accessibility and loss of fold architecture in mutant gelsolin at low pH as compared to WT gelsolin. Our findings support the existence of a secondary oligomerization-dependent aggregation pathway associated with gelsolin amyloidosis and can pave the way for better therapeutic strategies.

An Extended Guinier Analysis for Intrinsically Disordered Proteins

Guinier analysis allows model-free determination of the radius of gyration (Rg) of a biomolecule from X-ray or neutron scattering data, in the limit of very small scattering angles. Its range of validity is well understood for globular proteins, but is known to be more restricted for unfolded or intrinsically disordered proteins (IDPs). We have used ensembles of disordered structures from molecular dynamics simulations to investigate which structural properties cause deviations from the Guinier approximation at small scattering angles. We find that the deviation from the Guinier approximation is correlated with the polymer scaling exponent ν describing the unfolded ensemble. We therefore introduce an empirical, ν-dependent, higher-order correction term, to augment the standard Guinier analysis. We test the new fitting scheme using all-atom simulation data for several IDPs and experimental data for both an IDP and a destabilized mutant of a folded protein. In all cases tested, we achieve an accuracy of the inferred Rg within ∼3% of the true Rg. The method is straightforward to implement and extends the range of validity to a maximum qRg of ∼2 versus ∼1.1 for Guinier analysis. Compared with the Guinier or Debye approaches, our method allows data from wider angles with lower noise to be used to analyze scattering data accurately. In addition to Rg, our fitting scheme also yields estimates of the scaling exponent ν in excellent agreement with the reference ν determined from the underlying molecular ensemble.

Low-resolution SAXS and comparative modeling based structure analysis of endo-β-1,4-xylanase a family 10 glycoside hydrolase from Pseudopedobacter saltans comb. nov.

The structure and biophysical properties of endo β-1,4-xylanase (PsGH10A) of family 10 glycoside hydrolase were characterized. The modeled PsGH10A structure showed classical (β/α)8-barrel fold. Ramachandran plot displayed 99.1% residues in favored and 0.3% in the generously allowed region and only 0.6% residues in disallowed region. The secondary structure analysis of PsGH10A by CD revealed 31.75% α-helices 20.0% β-strands and 48.25% random coils. Protein melting study of PsGH10A showed complete unfolding at 60°C and did not require any metal ion for its stability. Structural superposition and docking analysis confirmed the involvement of Glu156 and Glu263 residues in catalysis. SAXS analysis displayed that PsGH10A is monomeric in nature showing fully folded state in solution form. Guinier analysis gave the radius of gyration (Rg) 2.23–2.29nm. Kratky plot indicated that the protein is fully folded globular shaped and flexible in solution form. The ab initio derived dummy model of PsGH10A displayed chicken thigh like shape. The ab initio derived dummy model superposed well with its comparative modeled structure except the N-terminal His-tag region.

SAXS and homology modelling based structure characterization of pectin methylesterase a family 8 carbohydrate esterase from Clostridium thermocellum ATCC 27405

Pectin methylesterase (CtPME) from Clostridium thermocellum of family 8 carbohydrate esterase (CE8) belongs to pectin methylesterase super family (E.C.3.1.1.11). BLAST analysis of CtPME showed 38% sequence identity with PME from Erwinia chrysanthemi. Multiple sequence alignment of CtPME with other known structures of pectin methylesterase revealed the conserved and semi-conserved amino acid residues. Homology modelling of CtPME structure revealed a characteristic right handed parallel β-helices. The energy of modelled structure was minimized by using YASARA software. The Ramachandran plot of CtPME shows 83.7% residues in non-glycine and non-proline residues in most-favorable region, 13.8% in additional allowed region and 1.4% in generously allowed region, indicating that CtPME has a stable conformation. The secondary structure of CtPME predicted using PSI-Pred software and confirmed by the circular dichroism (CD) showed α-helices (3.1%), β-sheets (40.1%) and random coils (56.9%). Small Angle X-ray Scattering (SAXS) analysis demonstrated the overall shape and structural characterization of CtPME in solution form. Guinier analysis gave the radius of gyration (Rg) 2.28 nm for globular shape and 0.74 nm for rod shape. Kratky plot gave the indication that protein is fully folded in solution. The ab initio derived dummy atom model of CtPME superposed well on modelled CtPME structure.

Angle-Dependent Light Scattering Study of Silica Aggregate Growth in 1-D Methane/Air Flames with Hexamethyldisiloxane Admixture: Effects of Siloxane Concentration, Flame Temperature, and Equivalence Ratio

ABSTRACTSilica aggregate formation was studied in 1D premixed methane/hexamethyldisiloxane/air flames by angle-dependent light scattering measurements for various siloxane concentrations, flame temperatures, and equivalence ratios, using Guinier analysis to interpret the experimental data. A sublinear dependence of the aggregate radii of gyration of generated silica particles on residence time, and non-monotonic dependence on flame temperature with a maximum around 2000 K have been observed, with radii of gyration in the range of 10 to 120 nm. Furthermore, a lean flame environment appears to foster aggregate growth compared to rich and stoichiometric flames, in which growth is very similar. When fixing the initial conditions at the residence time corresponding to the first measurement point, a simple model describing particle evolution as a result of collisional growth and sintering predicts well the functional dependence of the growth of particle radii.