Structure of native proteoglycan aggregates from chick limb-bud chondrocytes

Laser light-scattering has been used to investigate the size of native proteoglycan aggregates (PGA-aA1) from day-8 chick limb-bud chondrocyte cultures isolated under associative extraction and purification conditions in 0.4M guanidinium chloride (GdnHCl) solution. Dynamic light-scattering measurements yielded a hydrodynamic radius, R_s, of 244 ± 10 nm for PGA-aA1 in 0.4M GdnHCl, and a weight-average molecular weight (M _w) of 150 ± 50 × 10⁶ was obtained from a Zimm plot. Disaggregation in 4.0M GdnHCl aqueous solution yielded proteoglycan subunits (PGS) with R_s = 39 ± 2 nm, M _w = 1.6 ± 0.3 × 10⁶, which reassembled in 0.4M GdnHCl to form “reconstituted native” aggregates (PGA-raA1) with R_s = 121 ± 6 nm, M _w = 17 ± 3 × 10⁶. A second specimen of PGA-aA1 had R_s = 192 ± 10 nm, M _w = 100 ± 10 × 10⁶. The latter value was estimated from an empirical relationship between M _w and R_s. After dissociation, this specimen reassembled to form PGA-raA1 with R_s = 85 ± 5 nm, M _w = 12 ± 1 × 10⁶. These data are compared with those for a specimen of reconstituted aggregate (PGA-A1) that had been extracted under dissociative conditions and then reaggregated by dialysis to 0.4M GdnHCl aqueous solution, for which R_s = 138 ± 9 nm, M _w = 45 ± 8 × 10⁶. From these values, we have calculated the weight-average number of subunits per aggregate N_w: 111 for PGA-aA1 and 12 for raA1 (70 and 7 for the second PGA-aA1 and PGA-raA1 specimen, respectively) as compared to 32 for PGA-A1. The numbers of subunits per aggregate were also determined from electron micrographs of spread specimens. The latter results show the same trends as those obtained by light scattering, but lead in each case to lower numbers of subunits per aggregate. These data demonstrate conclusively that PGA samples exhibit a higher degree of aggregation in solution than visualized in typical electron microscopy (EM) preparations, probably due to disaggregation during EM specimen preparation. Since N_w determined both by light scattering (LS) and by EM are larger for native versus reconstituted aggregate samples, our data point to a more compact aggregation of subunits along the hyaluronic acid (HA) chains in the former.     

A New Preparation Method of Coenzyme A with Microorganisms

The structure of aggregates formed by heating dilute BSA solution was analyzed with the fractal concept using light scattering methods. BSA was dissolved in HEPES buffer of pH 7.0 and acetate buffer of pH 5.1 to 0.1% and 0.001% solutions, respectively, and heated at 95°C, varying the heating time t_a. The fractal dimension D_f of the aggregate in the solution was evaluated from static light scattering experiments. The polydispersity exponent τ and the average hydrodynamic radius <R_h> of the aggregates were calculated from dynamic light scattering experiments using master curves obtained by Klein et al. The values of D_f and τ of heat-induced aggregates of BSA at pH 7.0 were about 2.1 and 1.5, respectively, the values of which agreed with those predicted by the reaction-limited cluster–cluster aggregation (RLCCA) model. On the other hand, D_f of heat-induced aggregates at pH 5.1 was about 1.8, which agreed with that predicted by the diffusion-limited cluster–cluster aggregation (DLCCA) model. The dependence of <R_h> for the sample of pH 7.0 on t_a was similar to that of the polystyrene colloids reported previously.     

The molecular structure and solution conformation of an acidic heteropolysaccharide from Auricularia auricula-judae

A water soluble acidic heteropolysaccharide named WAF was isolated from Auricularia auricula‐judae by extracting with 0.9% NaCl solution. By using gas chromatography, gas chromatography‐mass spectrometry, and NMR, its chemical structure was determined to be composed of a backbone of α‐(1→3)‐linked D ‐mannopyranose residues with pendant side groups of β‐D ‐xylose, β‐D ‐glucose, or β‐D ‐glucuronic acid at position O6 or O2. Six fractions prepared from WAF with a weight‐average molecular mass (M_w) between 5.9 × 10⁴ and 64.7 × 10⁴ g/mol were characterized with laser light scattering and viscometry in 0.1M NaCl at 25°C. The dependence of intrinsic viscosity ([η]) and radius of gyration (R_g) on M_w for this polysaccharide were found to be [η] = 1.79 × 10^?3M_w^0.96 cm³ g^?1 and R_g = 6.99 × 10^?2 M_w^0.54 nm. The molar mass per unit contour length (M_L) and the persistence length (L_p) were estimated to be 1124 nm^?1 and 11 nm, respectively. The WAF exhibited a semirigid character typical of linear polysaccharides. Molecular modeling was then used to predict the ordered and disordered states of WAF; the simulated M_L and L_p were however much smaller than the experimental values. Taken altogether, the results suggested that WAF formed a duplex in solution. © 2010 Wiley Periodicals, Inc. Biopolymers 95: 217–227, 2011.     

Static and dynamic light scattering from dilute insulin solutions

Static and dynamic light-scattering measurements are reported on zinc-insulin at room temperature (21 ± l°C) and pH = 6.88 in 0.1M NaCl aqueous solution. The experiments were performed at very low concentration, in the range 0.12 × 10^?4 to 0.90 × 10^?4 g cm^?3. Within experimental error, we find no evidence for a critical micellar concentration in this system. The aggregation phenomenon starts immediately after preparation of the solutions, and takes several days to come to stable equilibrium. The concentration dependence of the diffusion coefficients, D _z, = D_o (1 — k_DC), is negative, and k_D was observed to decrease as a function of time, while the aggregate size was found to increase. The equivalent concentration coefficient, ?2BM _W, obtained from static light scattering, showed a similar behavior, and, within experimental error, was found to be numerically equal to k_D. From the relation found between the diffusion coefficient at infinite dilution and the molecular weight of the aggregates, log D₀ = ?0.240 log M _w ? 5.077, we deduce that the insulin aggregates are compact structures with a characteristic radius of 0.71 Å/(dalton)^1/3, surrounded by a hydration layer of a thickness of 8.0 Å. The equilibrium aggregation number is approximately 10.     

A complete hyaluronan hydrodynamic characterization using a size exclusion chromatography-triple detector array system during in vitro enzymatic degradation

Size exclusion chromatography coupled with triple detection (online laser light scattering, refractometry, and viscosimetry) (SEC-TDA) was applied for the study of hyaluronan (HA) fragments produced during hydrolysis catalyzed by bovine testicular hyaluronidase (BTH). The main advantage this approach provides is the complete hydrodynamic characterization without requiring further experiments. HA was hydrolyzed using several BTH amounts and for increasing incubation times. Fragments were characterized in terms of weight and number average molecular weights (M_w and M_n, respectively), polydispersity index (M_w/M_n), hydrodynamic radius (R_h), and intrinsic viscosity ([η]). The Mark-Houwink-Sakurada (MHS) curves (log [η] versus log M_w) were then derived directly. Fragments covering a whole range of M_w (10-900 kDa) and size (R_h = 4-81 nm) and presenting a rather narrow distribution of molar masses (M_w/M_n = 1.6-1.7) were produced. From the MHS curves, HA conformation resulted in a change from a random coil toward a rigid rod structure while decreasing the M_w. HA enzymatic hydrolysis in the presence of a BTH inhibitor was also monitored, revealing that inhibition profiles are affected by ionic strength. Finally, a comparison of the kinetic data derived from SEC-TDA with the data from rheological measurements suggested different strengths of the two methods in the determination of the depolymerization rate depending on the hydrolysis conditions.     

Characterization of group C meningococcal polysaccharide by light-scattering spectroscopy. III. Determination of molecular weight, radius of gyration, and translational diffusional coefficient.

Group-specific polysaccharides isolated by means of a cetavlon procedure are immunogenic in man and induce protective immunity against meningococcal meningitis. Minute quantities of the polymers in solution can act as vaccines. We now report the first characterization of a fractionated (C-1) group C polysaccharide in 0.4KM KCl and 0.05M sodium acetate by means of light-scattering spectroscopy. Independent measurements of refractive index increments, absolute scattered intensities, angular scattering intensities and line widths as a function of scattering angles and delay times at different concentrations using incident wavelengths of 632.8 nm from a He–Ne laser and of 488 nm from an argon–ion laser yield information on aggregation properties, molecular weight (M_r), radius of gyration 〈r⁰_g〉^1/2_z, translational diffusion coefficient 〈D〉⁰_z, and second virial coefficients A₂ and B₂ of C-1 polysaccharide. At relatively high ionic strength (0.04M KCl + 0.05M sodium acetate), we obtain for the C-1 polysaccharide in solution M_r = 5.15 × 10⁵, 〈r²_g〉^1/2_z = 345 Å, A₂ = 1.25 × 10^?4 ml/g, 〈D〉 = 1.16 × 10^?7 cm²/sec with a corresponding Stokes radius of 240 Å and B₂ = 4.4 ml/g. A₂ and B₂ are the second virial coefficients from intensity- and diffusion-coefficient measurements. The C-1 polysaccharide aggregates in solution and behaves hydrodynamically like random coils. Viscosity and sedimentation studies further confirm our conclusions that the fractioned C-1 polysaccharide aggregates in solution and EDTA can partially break up those aggregates. However, the system remains polydisperse even after adding an excess amount of EDTA. The weight-average molecular weight of the C-1 polysaccharide in solution depends upon ionic strength and exhibits a minimum at ～0.2M KCl. Finally, viscosity, light-scattering, and sedimentation results all show that the aggregated macromolecular system behaves like random-coiled polymers with no measurable shape factors.     

Filipin and its interaction with cholesterol in aqueous media studied using static and dynamic light scattering

Aggregation of filipin in aqueous medium and filipin-induced changes in cholesterol micelles have been studied using intensity and dynamic light scattering. The dependencies of filipin aggregate dimensions on concentration, solvent, and temperature were studied, and revealed that the aggregates do not have a well-defined geometry, i.e., a critical micelle concentration cannot be detected and stable structures are not formed. The aggregates are of size R_g ≈ 110 nm and R_h ≈ 63 nm, referring to the radius of gyration and hydrodynamic radius, respectively. In the concentration range studied (1 μM < C < 30 μM), a low molecular weight species (monomer/dimer) is always present together with the aggregates. In ethanol/ water mixtures, large (R_g ≈ 500 nm), narrow distribution aggregates are formed in the water volume fraction range 0.45 < Φ_H2_O < 0.65. Aggregation also occurs on changing the temperature; In the range 7–37°C, smaller aggregates (10–30 nm) form and the process is only partially reversible. No pronounced effect of filipin on the structure of the cholesterol micelles was observed (a small increase in R_g and R_h is noted). These results rule out any “specificity” for the filipin interactions with cholesterol, which has been considered a key event in the filipin biochemical mode of action. A reevaluatiori Of this question is suggested and some alternatives are advanced. © 1994 John Wiley & Sons, Inc.     

Molecular weight and pH aspects of the efficacy of oligochitosan against methicillin-resistant Staphylococcus aureus (MRSA)

Oligochitosan samples varying in molecular weight (M_w) and having narrow polydispersities were prepared by means of depolymerization of chitosan in hydrochloric acid, and their antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) was measured at pH values 5.5-8.0. The antibacterial testing of oligochitosans obtained showed that oligochitosans having M_w in the range of 0.73-20.0 kDa could be used both at slightly acidic and neutral pH values, and that the activity against MRSA remained moderate for oligochitosan samples having M_w about 3-5 kDa even at slightly basic pH values. The self-assembling behavior of oligochitosan macromolecules in the dilute solution at various pH values as a function of chain length was investigated. At first it was shown that oligochitosans formed supramolecular aggregates in dilute solutions below the critical pH value 6.5. Despite the aggregation phenomenon, the formation of nano-sized aggregates did not prevent oligochitosan from demonstrating the bactiostatic activity.     

Translational diffusion coefficient of α-chymotrypsinogen A by laser-light-scattering spectroscopy

The translational diffusion coefficient of a pure sample of α-chymotrypsinogen A is measured by laser light scattering to give a value of D_20,w⁰ = (8.40 ± 0.15) × 10^?7 cm²/sec.     

Physicochemical studies on xylinan (acetan). III. Hydrodynamic characterization by analytical ultracentrifugation and dynamic light scattering

A laboratory-made sample of the polysaccharide xylinan (acetan) has been further characterized with respect to (i) purity, (ii) molar mass and polydispersity, and (iii) gross conformation by a combination of hydrodynamic measurements (sedimentation velocity and equilibrium analytical ultracentrifugation, viscometry, and dynamic light scattering) in aqueous NaCl (I = 0.10 mol·L⁻¹). Sedimentation velocity diagrams recorded using Schlieren optics revealed highly pure material sedimenting as a single boundary [s^o_20.w = 9.5 ± 0.7) S; k_s = (273 ± 112) mL/g]. The hypersharp nature of these boundaries is symptomatic of a polydisperse and highly nonideal (in the thermodynamic sense) system. Low speed sedimentation equilibrium in the analytical ultracentrifuge using Rayleigh interference optics and two different types of extrapolation procedure (involving point and whole-cell molar masses) gave a weight average molar mass M_w of (2.5 ± 0.5) × 10⁻⁶ g·mol⁻¹ and also a second virial coefficient, B = (2.8 ± 0.7) × 10⁻⁴ mL·mol·g⁻², both values in good agreement with those from light scattering-based procedures (Part II of this series). A dynamic Zimm plot from dynamic light scattering measurements gave a z-average translational diffusion coefficient D^o_20.w = (3.02 ± 0.05) × 10⁻⁸ cm²·s⁻¹ and the concentration-dependence parameter k_D = (370 ± 15) mL/g. Combination of s^o_20.w with D^o_20.w via the Svedberg equation gave another estimate for M_w of ≅ 2.4 × 10⁶ g/mol, again in good agreement. Both the Wales-van Holde ratio (k_s/[η]) ≅ 0.4 (with [η] = (760 ± 77) mL/g) and the ρ-parameter (ratio of the radius of gyration from static light scattering to the hydrodynamic radius from dynamic light scattering) as ρ > 2.0 all indicate an extended conformation for the macromolecules in solution. These findings, plus Rinde-type simulations of the sedimentation equilibrium data are all consistent with the interpretation in terms of a unimodal wormlike coil model performed earlier. © 1996 John Wiley & Sons, Inc.     

Preparation and solution properties of pullulan fractions as standard samples for water-soluble polymers

A series of pullulan fractions with molecular weights in the range 5 × 10³ to 8 × 10⁵ were prepared. The weight-average molecular weight (M_w) of all the samples was determined by sedimentation equilibrium. The hydrodynamic properties of pullulan in aqueous solution were investigated by viscometry and ultracentrifugation. The experimental results indicate that pullulan molecules in water are fairly stable and behave as expanded random coils when M_w is above 2 × 10⁴. The molecular weight distributions of the fractions were measured by gel filtration. The ratio M_w/M_n was close to 1·1, except for a sample with the highest M_w.It is concluded that the pullulan fractions prepared by the present work are well characterized and have a narrow molecular weight distribution. They may be useful as standard samples for studies of water-soluble polymers.     

Analytical Ultracentrifugation Sedimentation Velocity for the Characterization of Detergent-Solubilized Membrane Proteins Ca++-ATPase and ExbB

We have investigated the potential of new methods of analysis of sedimentation velocity (SV) analytical ultracentrifugation (AUC) for the characterization of detergent-solubilized membrane proteins. We analyze the membrane proteins Ca⁺⁺-ATPase and ExbB solubilized with DDM (dodecyl-β-d-maltoside). SV is extremely well suited for characterizing sample heterogeneity. DDM micelles (s _20w?=?3.1 S) and complexes (Ca⁺⁺-ATPase: s _20w?=?7.3 S; ExbB: s _20w?=?4 S) are easily distinguished. Using different detergent and protein concentrations, SV does not detect any evidence of self-association for the two proteins. An estimate of bound detergent of 0.9 g/g for Ca⁺⁺-ATPase and 1.5 g/g for ExbB is obtained from the combined analysis of SV profiles obtained using absorbance and interference optics. Combining s _20w with values of the hydrodynamic radius, R _s?=?5.5 nm for Ca⁺⁺-ATPase or R _s?=?3.4 nm for ExbB, allows the determination of buoyant molar masses, M _b. In view of their M _b and composition, Ca⁺⁺-ATPase and ExbB are monomers in our experimental conditions. We conclude that one of the main advantages of SV versus other techniques is the possibility to ascertain the homogeneity of the samples and to focus on a given complex even in the presence of other impurities or aggregates. The relative rapidity of SV measurements also allows experiments on unstable samples.     

Structural analysis of purified human tracheobronchial mucins

Light scattering has been used to investigate the structure of human tracheobronchial mucin glycoproteins (HTBM) from the sputum of cystic fibrosis patients. The specimen was extracted using 6M guanidinium hydrochloride solution and fractionated by gel exclusion chromatography on Sephacryl S-1000. The fractionated HTBM was purified by density gradient ultracentrifugation. Purity of the resulting material was confirmed by SDS polyacrylamide gel electrophoresis and uv spectroscopy. Light scattering measurements on the fractionated mucins yield weight-average molecular weights M_w, and z-average radii of gyration R_{g, z}. The native cystic fibrosis HTBM consisted of a high molecular weight fraction with M_w = 9.3 × 10⁶ daltons and a lower molecular weight fraction contanining partly degraded mucins. After reduction and carboxymethylation of the high molecular weight native fraction, the resulting material was separated into three pools with M_w values of 5.1 × 10⁶, 1.6 × 10⁶, and 400,000. The derived molecular weights for the protein cores M_p,w, and the experimental radii of gyration are found to be consistent with the M_p,w – R_g relation established previously for submaxillary, cervical, and gastric mucins. These results imply that HTBM has the same extended-coil conformation reported for other mucins and has a molecular structure consisting of subunits, linked into linear chains via covalent (disulfide) bonds.     

Effect of water activity on the mechanical glass transition and dynamical transition of bacteria

The purpose of this study was to clarify the glass-transition behavior of bacteria (Cronobacter sakazakii) as a function of water activity (a_w). From the water sorption isotherm (298 K) for C. sakazakii, monolayer water content and monolayer a_w were determined to be 0.0724 g/g-dry matter and 0.252, respectively. Mechanical relaxation was investigated at 298 K. In a higher a_w range of over 0.529, the degree of mechanical relaxation increased with an increase in a_w. From the effect of a_w on the degree of mechanical relaxation, the mechanical a_wc (a_w at which mechanical glass transition occurs at 298 K) was determined to be 0.667. Mean-square displacement of atoms in the bacteria was investigated by incoherent elastic neutron scattering. The mean-square displacement increased gradually with an increase in temperature depending on the a_w of samples. From the linear fitting, two or three dynamical transition temperatures (low, middle, and high T_ds) were determined at each a_w. The low-T_d values (142–158 K) were almost independent from a_w. There was a minor effect of a_w on the middle T_d (214–234 K) except for the anhydrous sample (261 K). The high T_d (252–322 K) largely increased with the decrease in a_w. From the a_w dependence of the high T_d, the dynamical a_wc was determined to be 0.675, which was almost equivalent to the mechanical a_wc. The high T_d was assumed to be the glass-transition temperature (T_g), and anhydrous T_g was estimated to be 409 K. In addition, molecular relaxation time (τ) of the bacteria was calculated as a function of a_w. From the result, it is suggested that the progress of metabolism in the bacterial system requires a lower τ than approximately 6 × 10^?5 s.     

The flexibility of low molecular weight double-stranded dna as a function of length: I. Isolation and physical characterization of seven fractions

Sonicated calf thymus DNA was fractionated by rate zonal centrifugation into seven fractions with weight average molecular weights ranging from 0.28 to 1.3 × 10⁶ daltons, as determined by sedimentation equilibrium and light scattering measurements (the latter are described in the accompanying paper). Electron microscopy and sedimentation equilibrium analysis revealed these fractions to be narrowly disperse with M_w/M_n ratios averaging about 1.06. Intrinsic viscosities and sedimentation rates were measured and found to vary linearly with molecular weight in double-logarithmic plots in fair agreement with previously published functions relating these parameters for low molecular weight DNA. The average value for β from the Mandelkern— Flory equation was 2.59 × l0⁶, also agreeing with reported estimates of this parameter for short DNA. These data will be used in the second paper of this series to calculate the persistence length of the DNA fragments in each of the seven fractions by light scattering and hydrodynamic theories for the Kratky—Porod worm-like coil.     

Absolute molecular weight and molecular weight distribution of guar by size exclusion chromatography and low-angle laser light scattering

A procedure to determine the absolute weight-average molecular weight (M_w) and molecular weight distribution (MWD) of guar by aqueous size exclusion chromatography coupled with low angle laser scattering is described. It is shown that for a rigorously purified sample of guar solution the values for M_w and MWD are 2·2×10⁶ and 1·9 respectively. The effect of sample preparation and purification on these molecular parameters are discussed. Limitations and challenges in the aqueous size exclusion chromatography of complex water soluble polymers such as guar are also explored.     

Solution properties of porcine submaxillary mucin

Porcine submaxillary mucin (PSM) is a glycoprotein composed of a protein core and frequent, short oligosaccharide side chains. We report static and dynamic light scattering experiments and intrinsic viscosities for PSM in aqueous solvent systems. In 0.1M NaCl solution, the data suggest PSM exists as large, internally branched, highly hydrated, polydisperse aggregates that slowly dissociate to give a stable species of weight-average molecular weight (M_w) 7.4 × 10⁶. In 6M GdnHCl solution, the noncovalent bonds between PSM molecules are broken, giving a highly elongated molecule of M_w = 2.0 × 10⁶. The irreversible nature of this dissociation suggests that the forces that stabilize the native aggregates of PSM in 0.1M NaCl are specific in nature. On reduction of PSM with mercaptoethanol, the polydispersity decreases and M_w also decreases to 9 × 10⁵. A discrete change is observed in the solution properties of PSM in 0.1M NaCl at a concentration of 2mg/mL, manifested by a sudden decrease in the translational diffusion coefficient, an increase in viscosity number, and a decrease in slope of the osmotic compressibility. We tentatively propose that a weak and reversible secondary association process occurs at this concentration, although a purely hydrodynamic interaction cannot be ruled out.     

Chain conformation and bioactivity of water‐soluble polysaccharide extracted from Rhizoma Panacis Japonici

A water‐soluble α‐(1→4)‐D ‐glucan heteropolysaccharide with 37% degree of branch extracted by base from Rhizoma Panacis Japonici, coded as RPS3, was fractionated into six fractions by the method of nonsolvent addition. Their weight‐average molecular mass (M_w), polydispersity index (M_w/M_n), and radius of gyration (〈s²〉_z^1/2) were determined with laser light scattering (LLS) and size exclusion chromatography combined with LLS. The structure of the fraction was determined by methylation analyses and ¹³C NMR. The dependences of intrinsic viscosity ([η]) and 〈s²〉_z^1/2 on M_w were established as [η] = 0.71 M_w^{0.27 ± 0.01} (cm³/g) and 〈s²〉_z^1/2 = 1.53 M_w^{0.27 ± 0.02} (nm) in the M_w range from 5.62 × 10⁴ to 3.05 × 10⁶ (g/mol) for RPS3 in 0.15M NaCl aqueous solution at 25°C. On the basis of the current theory of the polymer solution, the fractal dimension (d_f), unperturbed chain dimension (A), and characteristic ratio (C_∞) were calculated to be 3.0, 1.48 Å, and 15.1, respectively. The results revealed that the RPS3 chains existed as spherical conformation in the aqueous solution. Transmission electron microscope further provided the evidence of the sphere shape of the RPS3 and its fractionated molecules in water. In vitro cytotoxicity assay indicated that the fractions could inhibit the tumor cells and showed no harm to normal cells at low dose. The bioactivity was relative with molecular mass of the samples. © 2010 Wiley Periodicals, Inc. Biopolymers 93: 383–390, 2010. This article was originally published online as an acceptedpreprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office atbiopolymers@wiley.com     

Light scattering during the hysteresis effect in poly(A)·2poly(U)

Light-scattering studies on buffered aqueous solutions of the triple-stranded polyribonucleic acid poly(A)·2poly(U) were carried out at neutral pH and during titration. At pH 7.1 and 22°C, a sample of commercially available polymer in 0.005M phosphate buffer gave a Zimm plot which yielded values for the weight-average molecular weight, M _w, of 874,000 ± 1800 g/mol, a root-mean-square radius, ρ of 930 ± 22 Å, and a second viral coefficient of 0.51 ± 0.05 × 10 ^?3 cm³g^?1 mol. The light-scattering data were also analyzed by serval linear and nonlinear least-squares programs which were devised to determine the model (e.g., rod, coil, or zigzag) which could best describe the shape of the molecule. It was found that a rodlike model, perhaps with a few bends, was in best overall agreement with the data. The assumption that the molecule is a thin rod leads to a value for the linear density of 206 g mol^?1 Å^?1 and a translation of 3.3 Å per residue. These values are also in close agreement with those expected for a triple-stranded, thin, base-stacked molecule. During titration from neutral pH with 0.1M HCl, the observed apparent molecular weight slowly increased until at about pH 3.5 a sudden, large increase (about 30-fold) occurred. The root-mean-square radius, on the other hand, after an initial small decrease (of about 25%), also exhibited a large increase (about 4-fold). Upon back titration with 0.1M NaOH, the molecular parameters did not retrace the original path, but instead exhibited hysteresis—the M _w and ρ _z are both larger on the basic branch than on the acid branch at a corresponding pH. A plot of long ρ _z against log(M _w) during the interval in which the high-moelcular-weight form was present (below pH 3.5 on the acid branch, and on the basic branch) gave a straight line with a slope of ?. This suggests that the aggregates were composed of some tens of rather open radom coils, presumably of poly(A)·poly(A), and that the hysteresis may be caused under conditions by the metastability of the entangled coils.