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1.
The cross-sectional radius of gyration of the deoxyribonucleoprotein (DNP) threads was measured by small-angle X-ray scattering in a wide range of ionic strengths (from 0.0005 to 2 M NaCl). For DNP in a solution of low ionic strength, this value is 30 Å. The increase of ionic strength results in partial deproteinization of DNP, while the cross-sectional radius of gyration varies from 25 Å for DNP in 0.7 M NaCl to 10 Å for DNP in 2 M NaCl. It is suggested that gradual deproteinization by the increase of NaCl concentration causes conformational changes, which are associated with the alteration of the DNP superstructure. The data are interpreted on the basis of the superhelical model of DNA packing in DNP; however, the coexistence of superhelical and unfolded regions in the DNP structure is also a possibility.  相似文献   

2.
M Boublik  N Brot  H Weissbach 《Biopolymers》1973,12(9):2083-2092
Ribosomal proteins L7 and L12 are the only acidic proteins found on the 50S ribosomal subunit of Escherichia coli. The effect of ionic strength, helix-promoting solvents and denaturating agents on the conformation of these proteins has been studied. It has been established that the helicity of L7 and L12 proteins (approx. 45–50% α helix) can be increased to 60–70% when they are exposed to helix-promoting solvents such as methanol or ethanol in the presence of 0.1M salt. High ionic strength by itself was without any effect on the conformation of the proteins. However, the solvent, 2,2,2-trifluoroethanol increased the content of α helices up to 80% even in the absence of salt. Denaturating agents like urea (6M) or guanidine HCl (6M), decreased the content of the ordered structure below 20%. All conformational changes induced by salt or solvents were completely reversible and characterized by a broad transition showing a low degree of cooperativity. This might indicate the presence of discrete segments with variations in amino acid sequences and ordered structures with different stabilities.  相似文献   

3.
The secondary structure of xanthan in solutions of relatively low salt concentration and at room temperature has been investigated using static light scattering experiments. Additional evidence has been found for a dimeric structure at 25°C in 0.01M NaCl. From the experimental z-average mean square (ms) radius of gyration, a value for the persistence length p has been estimated, taking explicitly into account the polydispersity of the three samples used, which has been established by gel permeation chromatography (GPC) measurements. The experimental particle scattering functions of the three samples are consistent with theoretical estimates for polydisperse systems with the same value of p = 65 ± 10 nm and the molar mass per unit length for a dimeric structure. This secondary structure remains unaffected by the ionic strength in the 0.005–0.0lM range. Partial aggregation seems to occur at higher NaCl concentrations. Light scattering and GPC data show that heating the xanthan 0.01M NaCl solutions to about 70°C considerably reduces the Mw of the low molar mass sample (2.3 × 105-g·mol?1), contrary to what is observed for the high molar mass sample (1.8 × 106-g·mol?1). These experimental findings can be accounted for by a partial temperature-induced dissociation of the xanthan dimers according to an all-or-none mechanism. © 1994 John Wiley & Sons, Inc.  相似文献   

4.
Recent studies on dilute solutions (Borgström et al. (1996), Int. J. Biol. Macromol. 18, 223) have shown that kappa-carrageenan helices associate into superhelical rigid rods in mixed 0.1 M aqueous solutions of NaI and CsI above a critical mole fraction (xCs = 0.4) of Cs. This work concerns the temperature-dependent rheology of more concentrated systems in mixed and pure solutions of the same salts. Gels with low moduli were even found in NaI alone, although this salt is known to impede the gelation of kappa-carrageenan, but only above 0.9% (w/w) of carrageenan. These gels were reminiscent of iota-carrageenan gels in two respects: the (low) magnitude of the shear storage modulus (G′), and the absence of hysteresis in the sol-gel transition. On the other hand, both the threshold concentration for gelation and the ratio between the loss and storage moduli were substantially higher for the kappa-carrageenan gels in NaI. In mixed solutions of CsI and NaI, two types of kappa-carrageenan gels could be distinguished, depending on the cesium content. The transition occurred at xCs = 0.4, as in the previous studies on dilute solutions. Below xCs = 0.4, the gels were similar to those in NaI alone. Above xCs = 0.4, the gels were similar to ‘conventional’ kappa-carrageenan gels, formed in salts such as KC1: a pronounced thermal hysteresis appeared in the sol-gel transition, the gels showed tendencies for syneresis, and G′ increased dramatically with increasing cesium content.  相似文献   

5.
The viscosity of an exocellular polysaccharide (EPS) produced by the bacterium Lactococcus lactis subsp. cremoris B40 was studied in aqueous solution at an ionic strength of 0.10M. First, the zero‐shear viscosity was determined as a function of the concentration. From the data in the low concentration range, the intrinsic viscosity was determined. In addition, the shear‐thinning behavior was measured at several concentrations. By combining existing theories, a new equation is proposed that describes and predicts the intrinsic viscosity and the concentration dependence of the (zero‐shear) viscosity of B40 EPS solutions from the molar mass and the hydrodynamic radius of the polysaccharide. Based on the Rouse theory, the shear‐rate dependence of the viscosity also could be described and predicted from the molecular characteristics, i.e., molar mass and radius of gyration. It is shown that these equations can be applied to all random coil polysaccharides. © 1999 John Wiley & Sons, Inc. Biopoly 50: 641–646, 1999  相似文献   

6.
Isothermal microcalorimetry, differential scanning calorimetry (DSC), and chirooptical data obtained for ι-carrageenan in NaCl, LiCl, and NaI aqueous solutions are presented. The experiments have been performed as a function of concentration both for the polymer and for the simple salt as a cosolute. The experimental findings consistently show the occurrence of a salt-induced disorder-to-order transition. From microcalorimetric experiments the exothermic enthalpy of transition ΔHtr is obtained as the difference between the theoretical, purely electrostatic ΔHel enthalpy change and the actual mixing enthalpy ΔHmix, measured when a ι-carrageenan salt-free solution at constant polymer concentration is mixed with a 1:1 electrolyte solution of variable concentration. In the case of added NaCl, the absolute values of enthalpy changes |ΔHtr| are in good agreement with those obtained for the opposite process, at comparable polymer and salt concentrations, from DSC melting curves. The microcalorimetric results show that the negative maximum value of ΔHtr corresponding to the interaction of Li+ counterion with ι-carrageenan polyion results to be significantly lower than the corresponding values obtained for Na+ counterion. At variance with the microcalorimetric data, chirooptical results show that the salt-induced disorder-to-order transition, occurring in the 0.02–0.2M salt concentration range, appears to be complete at a concentration of about 0.08–0.1M of the simple ion, irrespective of the polymer concentration and of the nature of added electrolyte. © 1998 John Wiley & Sons, Inc. Biopoly 45: 105–117, 1998  相似文献   

7.
The structure of human plasma fibronectin in 50 mM Tris-HCl buffer, pH 7.4, containing varying concentrations of NaCl, has been investigated using the small-angle X-ray method.Below 0.3 M NaCl the overall structure of the molecule is disc-shaped; at 0 M NaCl the axial ratio of the disc is about 1:7 and between 0.1 M to 0.3 M it is slightly more asymmetric, with an axial ratio of 1:10.At about 0.3 M NaCl there is a reversible transition to a more open structure, and, from 0.3 M up to 1.1 M NaCl the small-angle X-ray data can be explained by models consisting of ensembles of flexible, non-overlapping, bead-chains generated by a Monte Carlo procedure. Within this concentration range there is a gradual increase in the stiffness of the chains, as well as a decrease in bead radius, which indicates that the molecule becomes more open when the NaCl concentration is increased.The transition to a more open structure is also demonstrated by the average radius of gyration which increases gradually from 8.26 nm at 0 M NaCl to 8.75 nm at physiological or near-physiological conditions, and up to 16.2 nm at 1.1 M NaCl.Abbreviations hpFN human plasma fibronectin - SAXS smallangle X-ray scattering - Tris tris (hydroxymethyl) aminomethane - DTT dithiothreitol - BA benzamidine hydrochloride - PMSF phenylmethylsulfonyl fluoride  相似文献   

8.
Protein tyrosine nitration is well-established post-translational modification occurring in a number of diseases, viz. neurodegenerative, cardiovascular diseases, ageing, etc. Tyrosine-67 (Tyr-67) nitration of cytochrome c (cyt c) was observed under oxidative stress affecting its structure and electron transfer properties. Hence, in this study, molecular dynamics (MD) simulations were carried out at room temperature to investigate the structural and conformational changes in the nitrated cyt c's. MD results revealed that the bond between FE (Heme-105) and S (Met-80) considerably weakened, radius of gyration, backbone and Cα root-mean-square deviations decreased and hydrogen bonding increased in the nitrated cyt c's relative to wild type (WT) cyt c. Ramachandran plot analysis revealed that N- and C-terminal helices also affected by nitration at CE2 carbon atom. Furthermore, essential dynamics analysis showed that amplitude of concerted motion decreased in the nitrated cyt c's, perhaps due to the increase in the hydrogen bonding interaction. Taken together, the structural and conformational changes in the active site Tyr-67 nitrated cyt c may have implications in the loss of electron/proton transfer and gain of apoptotic properties.  相似文献   

9.
The conductivity of xanthan (extracellular polysaccharide from Xanthomonas campestris) in the potassium salt form has been studied over the temperature range 5–80°C spanning the order-disorder conformational transition. In salt-free solution data analysis using Manning's polyelectrolyte-conductivity theory gives a charge spacing, b, of 0.58±0.04 nm for the low temperature ordered form, consistent with a single rather than a double helix (b=0.58 and 0.29 nm respectively). In solutions with 0.01 M added KBr the increase in counterion condensation on conformational ordering is found from conductivity studies to be — –1= 0.20 ± 0.02, in good agreement with the value 0.20±0.02 using polyelectrolyte-equilibrium theory for the variation of transition-midpoint temperature with added salt determined from opticalrotation data.  相似文献   

10.
Mixtures of locust bean gum (LBG) with κ-carrageenan (KC) in 0.1 M aqueous solutions of the mixed salts NaI/CsI were investigated by cryo-transmission electron microscopy (cryo-TEM) and dynamic viscoelastic measurements. Previous studies have shown that as the cesium content is increased in such mixed salt solutions, a transition occurs from molecularly dispersed helices to ‘superhelical rods’ of KC. We now found that LBG stabilises the superhelical rods, shifting the transition to a lower content of Cs for the mixtures than for KC alone. The formation of superhelical rods was evidenced both by cryo-TEM images and by an onset of thermal hysteresis in the coil–helix transition of KC. In the mixtures, the transition temperatures on cooling and heating were insensitive to the proportions of LBG and KC present at all cesium contents. Under conditions where no helix aggregation occurred (no hysteresis) the mixtures showed high tan δ values and low storage moduli. Under aggregated conditions, gels formed, and gels with added LBG had enhanced moduli compared to gels with KC alone. On the basis of these results we propose that LBG associates to the super-helical rods of KC.  相似文献   

11.
Candida antarctica lipase B (CALB), a serine protease, is involved in the hydrolysis of substrates at the aqueous lipid interface. There is a significant role played by the helices in serine proteases including acting as a flap covering the active site region. The α5 and α10 helices in the path to the active site of CALB appear to play an important role in the region. This study investigates these helices by mutational studies, docking and molecular dynamics simulations. The mutations were selected based on their proximity to the active site and their presence at the α10-helix in the path of the active site. Molecular dynamics studies reveal the flexibility, stability and hydrogen bonding ability of the α5 helix. The radius of gyration (R g) clearly showed the compactness of the structure. Docking studies show the changes occurring at the protein's binding site before and after 15 ns of simulation. Results from the study demonstrate the importance of the two helices α5 and α10 in the stability of CALB.  相似文献   

12.
N Murai  S Sugai 《Biopolymers》1974,13(6):1195-1203
The conformational changes of poly-Nε-glutaryl-L -lysine (PGL) and poly-Nε-succinyl-L -lysine (PSL) in various salt solutions were studied by use of ORD and potentiometric titration measurements. The addition of alkali metal salts to the fully ionized PGL or PSL solution caused helix formation. The helical content of the polymers increases in the following sequences: at salt concentration 0–2 M, CsCl < KCl < LiCl < NaCl; and at 2–3 M, LiCl < CsCl < KCl ~ NaCl. The preferential binding of the solvent components with various alkali metal salts of PGL or PSL was measured in LiCl, NaCl, and KCl solutions by means of equilibrium dialysis and differential refractometry. It was found that with increasing salt concentration, the polymers were preferentially hydrated in NaCl and KCl soultions; however the salt was preferentially bound to the polymers in LiCl solution. Such preferential binding was suggested to be closely related to conformational change. The addition of CaCl2 to polymer solutions led to the stabilization of the helical structure of PGL or PSL.  相似文献   

13.
Thermally induced order-disorder conformational transition in succinoglycan was studied using the method of high-sensitivity differential scanning microcalorimetry within the range of polysaccharide concentrations from 0.1 to 3.5 mg mL−1 at NaCl concentrations 0, 0.01, and 0.1M. The positions and shapes of the excess heat capacity curves depended substantially on both the NaCl and polysaccharide concentrations. At low polysaccharide concentrations in salt-free solution the experimental curves were closely approximated by the two-state model suggesting the transition mechanism to be of the single helix-coil type. With increasing polysaccharide and/or NaCl concentration, the experimental curves changed significantly in symmetry, which indicated a changing transition mechanism. At high polysaccharide concentrations or in the presence of the salt, the order-disorder transition of succinoglycan was shown to include two stages: the cooperative dissociation of the helix dimer and subsequent two-state melting of the helix monomer. The dependence of thermodynamic parameters for the dissociation and melting of helix structures in succinoglycan on NaCl and polysaccharide concentrations was obtained by fitting the experimental excess heat capacity curves. The cooperativity parameter σ for the single helix-coil transition as well as the average length of the helix segment of succinoglycan were calculated. Some features of succinoglycan ordering in solution are discussed. © 1996 John Wiley & Sons, Inc.  相似文献   

14.
The proteoglycan subunit (PGS) from bovine nasal cartilage was examined in water and in 0.15 N LiCl by small-angle x-ray scattering (SAXS). The molecular weight of 2.5 × 106 and the radius of gyration, Rg = 493 Å, in 0.15 N LiCl, obtained by SAXS, are in good agreement with values reported by others for similar preparations. Values of the radius of gyration of the cross section, mass per unit length, and persistence length of the PGS are also reported. The low value of intrinsic viscosity ([η]) found in 0.15 N LiCl, and a comparison of the experimental distance distribution function to that of the theoretical distance distribution function for sphere, suggest that the PGS in salt solution approaches spherical symmetry. The much higher value of [η] in water suggests a prolate ellipsoid of low axial ratio.  相似文献   

15.
Density (rho), apparent molar volume (V(phi)), and viscosity (eta) of 0.0010 to 0.0018% (w/v) of bovine serum albumin (BSA), egg albumin, and lysozyme in 0.0002, 0.0004, and 0.0008 M aqueous RbI and CsI, and (dodecyl)(trimethyl)ammonium bromide (DTAB) solutions were obtained. The experimental data were regressed against composition, and constants are used to elucidate the conformational changes in protein molecules. With salt concentration, the density of proteins is found to decrease, and the order of the effect of additives on density is observed as CsI > RbI > DTAB. The trend of apparent molar volume of proteins is found as BSA > egg-albumin > lysozyme for three additives. In general, eta values of BSA remain higher for all compositions of RbI than that of egg-albumin for CsI and DTAB. These orders of the data indicate the strength of intermolecular forces between proteins and salts, and are helpful for understanding the denaturation of proteins.  相似文献   

16.
For many protein therapeutics including monoclonal antibodies, aggregate removal process can be complex and challenging. We evaluated two different process analytical technology (PAT) applications that couple a purification unit performing preparative hydrophobic interaction chromatography (HIC) to a multi-angle light scattering (MALS) system. Using first principle measurements, the MALS detector calculates weight-average molar mass, Mw and can control aggregate levels in purification. The first application uses an in-line MALS to send start/stop fractionation trigger signals directly to the purification unit when preset Mw criteria are met or unmet. This occurs in real-time and eliminates the need for analysis after purification. The second application uses on-line ultra-high performance size-exclusion liquid chromatography to sample from the purification stream, separating the mAb species and confirming their Mw using a µMALS detector. The percent dimer (1.5%) determined by the on-line method is in agreement with the data from the in-line application (Mw increase of approximately 2750 Da). The novel HIC-MALS systems demonstrated here can be used as a powerful tool for real-time aggregate monitoring and control during biologics purification enabling future real time release of biotherapeutics.  相似文献   

17.
We show that the persistence length a of DNA, derived from total intensity laser light scattering of linear Col E1 DNA and corrected for excluded-volume effects, varies from about 68 nm in 0.005M NaCl to about 40 nm in 0.2M NaCl, leveling off to a constant value (about 27 nm) at high NaCl (1–4M) concentration. These observations do not agree with current views on the effect of electrostatic charge and ionic conditions on DNA dimensions. The apparent diffusion constant Dapp, determined from laser light scattering autocorrelation as a function of scattering vector q, at NaCl concentrations 0.005–4M, correctly yields the translational diffusion coefficient Dt at low values of q and scales with molecular dimensions rather than segment length at high values of q; thus, Dapp/Dt yields a universal curve when plotted against q2R, where Rg is the radius of the gyration. The sedimentation coefficients s at 0.1 and 0.2M NaCl concentration closely agree with the well-tested empirical relations, and a combination of s, Dt, and the appropriate density increments yield correct molar masses over the whole salt concentration range. Approximate constancy of DtRg indicates limited draining in translational flow. We present some observations and thoughts on the regimes in which a dependence of the correlation decay times on q3 rather than q2 applies. We conclude that quasielastic laser light scattering discloses little information about dynamics of internal motion of DNA chains.  相似文献   

18.
The hydrodynamic characteristics of heparin fractions in a 0.2 M NaCl solution have been determined. Experimental values varied over the following ranges: the sedimentation coefficient (at 20.0 °C), 1.3<s0×1013<3.2 s; the Gralen coefficient (sedimentation concentration-dependence parameter), 10<ks<70 cm3 g–1; the translational diffusion coefficient, 3.9<D0×107<15.4 cm2 s–1; the intrinsic viscosity, 7.9<[]<40 cm3 g–1. Combination of s0 with D0 using the Svedberg equation yielded molecular weights in the range 3.9<M×10–3<37 g mol–1. The value of the mass per unit length of the heparin molecule, ML, was determined using the theory of hydrodynamic properties of a weakly bending rod, giving ML=570±50 g nm–1 mol–1. The equilibrium rigidity, Kuhn segment length (A=9±2 nm) and hydrodynamic diameter (d=0.9±0.1 nm) of heparin were evaluated on the basis of the worm-like coil theory without the excluded volume effect, using the combination of hydrodynamic data obtained from fractions of different sizes. Small-angle X-ray scattering for three heparin fractions allowed an estimate for the cross-sectional radius of gyration as 0.43 nm; from the evolution with the macromolecule contour length of the radius of gyration, a value for the Kuhn segment length of 9±1 nm was obtained. A good correlation is thus observed for the conformational parameters of heparin from hydrodynamic and X-ray scattering data. These values describe heparin as a semi-rigid polymer, with an equilibrium rigidity that is essentially determined by a structural component, the electrostatic contribution being negligible in 0.2 M NaCl.Presented at the conference for Advances in Analytical Ultracentrifugation and Hydrodynamics, 8–11 June 2002, Grenoble, France  相似文献   

19.
Rigidity of myosin and myosin rod by electric birefringence   总被引:1,自引:0,他引:1  
S Hvidt  T Chang  H Yu 《Biopolymers》1984,23(7):1283-1294
The rotational relaxation times of rabbit myosin and myosin rod have been determined by electric birefringence measurement. The relaxation time of myosin measured in 10 mM pyrophosphate buffers in a pH range of 7.6–9.5 was found to have substantial concentration and pH dependences. The infinite-dilution limit of the relaxation time, τ°, was determined as 38 ± 2 μs, and it was found to be independent of pH. For myosin rod, a possible thermally induced conformational change was investigated in a temperature range of 1–43°C. The rotational relaxation time of myosin rod shows no clear indication of conformational change in this temperature range, and the radius of gyration measurement by light scattering was shown to be consistent with this observation. The steady-state birefringence, however, decreases substantially above around 40°C. This, the myosin rod appears to be only slightly flexible even at physiological temperature, but the possibility of a “melting” or “hinging” of the myosin rod cannot completely be ruled out on the basis of these experiments.  相似文献   

20.
Conformation of mucous glycoproteins in aqueous solvents   总被引:5,自引:0,他引:5  
Light-scattering techniques have been used to measure the z-average radius of gyration Rg z-average translational diffusion coefficient Dt and weight–average molecular weight Mw of porcine submaxillary mucin (PSM) in solution. PSM isolated at low shear in the presence of protease inhibitors has a Mw about twice as large as a sample prepared without these precautions. The former sample has a Mw of 17 × 106 in 0.1M NaCl, which decreases to 8 × 106 in 6M guanidine hydrochloride (GdnHCl) and then to 2 × 106 on addition of 0.1M mercaptoethanol to the 6M GdnHCl solution. The Rg or D values obtained for PSM in this work superimpose with those of other authors for different mucin glycoproteins, leading to linear log–log relationships to the molecular weight of the protein core. Comparison of these results with those in the literature for denatured proteins suggest that mucins are linear random coils in which the protein core is stiffened by the presence of the oligosaccharide side chains. The length of the oligosaccharides and the nature of the solvent have little effect on the extension of the protein core. This suggests that the stiffness of the protein core is maintained by steric repulsion of the residues at the beginning of the oligosaccharide chains.  相似文献   

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