The conformation and aggregation of bovine β-casein A. II. Thermodynamics of thermal association and the effects of changes in polar and apolar interactions on micellization

A sedimentation analysis has been used to determine the proportion of protein present as monomer and aggregate in 0.5 and 1.0 g/dl solutions of β-casein A in pH 7 phosphate buffer over the temperature range 10–40°C. The amount and molecular weight of the aggregate increase with temperature; under the conditions used, the aggregation number (n) of β-casein is given approximately by n = 0.6t + 2 with t in degrees centigrade. The concentration of β-casein in monomeric and aggregated states at different temperatures is used to calculate the standard enthalpy of aggregation ΔH° (Van't Hoff) by assuming that β-casein undergoes a cooperative, two-state, micellization process; aggregation is an endothermic process and ΔH° = 66.0 ± 2.6 kJ mol^?1. Combination of this ΔH° with the amount of protein calculated to dissociate when 1 g/dl solutions are diluted isothermally to 0.5 g/dl gives the heat of dilution at various temperatures. These calculated heats of dilution are compared with the experimental values obtained by carrying out the same dilutions in a microcalorimeter. The heat of dilution decreases linearly with β-casein concentration, but the extrapolated zero-concentration values of 65.8 ± 1.6 kJ mol^?1 is the same as the Van't Hoff enthalpy. This agreement in the enthalpy values indicates that the micellization of β-casein occurs cooperatively. The effect of modifying the hydrophobic/hydrophilic balance of the system on the micellization of β-casein A has been investigated. The hydrophobic interaction between the protein molecules is decreased by removing the three C-terminal residues (Ileu Ileu Val) with carboxypeptidase-A. This modification drastically reduces the ability of the β-casein molecule to form micelles. Substitution of ²H₂O for H₂O at constant temperature perturbs the monomer–micelle equilibrium in favor of micelles because of enhanced hydrophobic interactions in the former solvent. The results are consistent with β-casein micellization involving a delicate balance of the hydrophobic forces favoring aggregation and electrostatic forces opposing it.     

Melanostatin conformations in solution.

The conformations of melanostatin have been studied experimentally using CD spectroscopy and via calculations. In aqueous solution and 2,2,2-trifluoroethanol (TFE) there is no evidence that monomers of the tripeptide exist in an ordered (β-bend) structure. In water and TFE solutions (3–6 × 10^?4M) the neutral molecules aggregate very slowly, taking about 3 days to attain equilibrium at room temperature. At equivalent concentrations in TFE, although not in water, the cationic molecules also slowly aggregate, although to a lesser extent. Calculations using rotational isomeric state theory give the most probable unperturbed end-to-end distance of the molecule at 9.3 ± 0.1 Å and indicate that a vast majority of the molecules exist in some extended conformation, end-to-end distance ≥6 Å. Only 0.4% of the molecules are calculated to have O…?H separations compatible with a β-bend structure. An intramolecular hydrogen bond must have an energy at least 2 kcal/mol lower than that of an intermolecular hydrogen bond to solvent if a β-bend is to be experimentally observable.     

Comparison of conformations of κ-casein,para-κ-casein and glycomacropeptide

The conformation of κ-casein was compared with those of para-κ-casein and glycomacropeptide formed by the cleavage of κ-casein with chymosin. Para-κ-casein is insoluble in water at room temperature, but is slightly soluble in 0.07 M NaCl (pH 7.0) at 3°C. The secondary structure of κ-casein, para-κ-casein and glycomacropeptide was estimated from CD spectra measured at 3°C by the method of Yada and that of Provencher and Glockner. The surface hydrophobicity of these molecules was estimated by the fluorometric method. It was concluded that the secondary structures of para-κ-casein and glycomacropeptide segments were changed slightly by cleavage with chymosin. Para-κ-casein was estimated to have more β-sheet structure than glycomacropeptide. Para-κ-casein had larger hydrophobic regions on the molecular surface compared with the corresponding part in κ-casein.     

The conformation of polyriboadenylic acid at low temperature and neutral pH. A single-stranded rodlike structure.

The conformation of single-stranded polyrA in aqueous solution has been measured at temperatures down to ?12°C. The radius of gyration of low-molecular-weight polyrA varies very little with temperature in this range. By studying the dependence of radius of gyration on temperature for several polyrA fractions, we show that the dependence of the radius upon chain length is consistent with formation of a single-stranded rodlike structure at low temperature. The structure has an approximate length of 3.2 Å/nucleotide.     

Small-angle x-ray scattering by poly-L-alanine solutions

The radius of gyration and “persistence length” of poly-L -alanine, calculated from small-angle x-ray scattering data, have values of 56 Å and 44 Å, respectively, in dichloroacetic acid, and 78 Å and ～30 Å in a 1:1 v/v mixture of trifluoroacetic acid and trifluoroethanol. This can be interpreted to mean that poly-L -alanine exists in a relatively rigid, predominantly α-helical conformation in dichloroacetic acid and in an extended, more flexible form in the mixed solvent system.     

Theoretical Study of the Conformations of Pustulan [(1⇒6)-β-D)-Glucan]

Abstract

The total potential energy including nonbondedJuntorsional and hydrogen bond contributions has been computed for pustulan, a (1?6) linked β-D-glucan polysaccharide, as a function of rotational angles φ, ψ, and ω The (φ, ψ, ω)-space contains many local minima and at least three distinct deep minima. Two minima at (φ, ψ, ω)=(25°,190°,gg) and (φ, ψ, ω)=(65°,150°,gg) of almost equal energies have helical parameters (n=5.2, A=1.0Å) and (n=3.2, h= 1.5Å), respectively. A third minimum at (φ, ψ, ω)=(40°,70°gt) leads to an extended zig-zag structure (n=2.2, h=2.2Å). Energy maps obtained for gentiobiose, the disaccharide of pustulan, also reveal many local minima and the small energy differences among them indicate that gentiobiose is extremely flexible. Gentiodextrins, a family of cyclic molecules of (l?6)-β-D- glucose residues, were also studied. Conformations free from steric hindrance were found for cyclic molecules with three to six glucose residues.     

Crystalline structure of the gas vesicle wall from Anabaena flos-aquae.

The gas vesicles isolated from Anabaena flos-aquae have been studied by X-ray diffraction. Electron microscopy has previously shown that the gas vesicles are elongated shapes, with a thin wall having regular striations (ribs) at right-angles to the long axis. The X-ray diffraction pattern from a specimen of oriented, intact vesicles includes a number of sharp reflections which are attributed to regular structure in the plane of the wall. After correcting for the imperfect alignment of the long axes of the vesicles, the in-plane reflections are all seen to lie on a few, regularly spaced lines parallel to the long axis. This result shows for the first time that there are subunits regularly spaced along each rib, one subunit every 11 Å. The spacing of the in-plane reflections along each line is consistent with a rib periodicity of 46 Å. The 11 Å repeat, together with the 46 Å repeating distance from rib to rib and the average wall thickness of about 20 Å, define a volume for the subunit. Assuming a reasonable value for the density of the protein making up the wall, the molecular weight of the subunit indicated is about 8000 g/mol.The X-ray data also indicate that a large part of the protein is in the β-sheet conformation. In this structure there are parallel, or anti-parallel, polypeptide chains which are hydrogen-bonded to one another in a regular way to form a thin sheet. Assuming the wall contains β-sheet in two layers, one on top of the other and with the chains in each layer tilted at 35 ° to the long axis of the vesicle, we can explain a number of the X-ray observations: (1) oriented arcs with a Bragg spacing of 4.7 Å, which is the distance between the axes of neighbouring chains in each layer; (2) diffraction oriented in the direction of the chains at a spacing of 6 to 7 Å, which is the repeating distance of the dipeptide unit along the chain; (3) the 11 Å repeat, which is the repeating distance of pairs of chains along each rib; and (4) a broad band of diffraction at right-angles to the plane of the wall and centred at a spacing of 10 Å, which is a reasonable value for the distance between the mid-planes of the two sheets. Moreover, we can also find the remaining lattice parameter, the angle relating the centres of the subunits in neighbouring ribs. Thus the shortest line joining the centres makes an angle of 86 ° with the direction of the ribs.     

X-ray studies on crystalline complexes involving amino acids and peptides. XXIV. Different ionization states and novel aggregation patterns in the complexes of succinic acid with DL-and L-histidine

Diffusion of acetonitrile into an aqueous solution of DL -histidine and succinic acid in 1:3 molar proportions results in the crystals of DL -histidine hemisuccinate dihydrate [triclinic, P1 , a = 7.654(1), b = 8.723(1), c = 9.260(1) Å, α = 77.23(1), β = 72.37(1) and γ = 82.32 (1)°]. The replacement of DL -histidine by L -histidine in the crystallization experiment under identical conditions leads to crystals of L -histidine semisuccinate trihydrate [orthorhombic, P2₁2₁2₁, a = 7.030 (1), b = 8.773 (1), and c = 24.332 (3) Å]. The structures were solved using counter data and refined to R values of 0.056 and 0.054 for 2356 and 1778 observed reflections, respectively. Histidine molecules in both the complexes exist in open conformation I. Succinate and semisuccinate ions in them are planar, and exactly or nearly centrosymmetric. In the DL -histidine complex, the amino acid molecules form double ribbons and the succinate ions occupy voids left behind when the double ribbons aggregate, as in inclusion compounds. In the L -histidine complex, the amino acid molecules form columns; so do the semisuccinate ions and water molecules. The two columns interdigitate to form the complex crystal. There are similarities between the molecular aggregation in the complexes and that in the crystals of L - and DL -histidine. However, the presence of succinic acid has the effect of disrupting, partially or totally, head-to-tail sequences involving amino acid molecules. © 1993 John Wiley & Sons, Inc.     

Self-association of β-lactoglobulin A in acid solution. I. Translational diffusion coefficients

We report measurements of the diffusion coefficient of β-lactoglobulin A (βLG-A) at pH = 5.60 and 4.58 in 0.10 ionic strength acetate buffer by the techniques of analog photocurrent signal correlation and digital single-clipped photon correlation. At a concentration of 21 mg/ml and a pH of 4.58, the self-association of β-lactoglobulin can be represented by a simple dimer–octamer equilibrium model. We determined the translational diffusion coefficient of the dimer and that of the octamer using the scattering results of Kumosinski and Timasheff in a dimer–octamer mixture. Our analysis shows that the dimer βLG-A does not change its size if the pH is varied from 5.60 to 4.58 and both species remain constant in size for temperature changes from 3.5° to 25°C Hydrodynamically, the octamers behave like closed-packed spheres with an effective radius of about 45 Å according to the Stokes-Einstein relation.     

Effects of End Group and Aggregation on Helix Conformation: Crystal Structure of Ac-(Aib-Val-Ala-Leu)2-Aib- OMe

The role of end groups in determining stereochemistry and packing in hydrophobic helical peptides has been investigated using an α-aminosobutyric acid (Aib) containing model nonapeptide sequence. In contrast to the Boc-analogue, Ac-(Aib-Val-Ala-Leu)₂-Aib-OMe crystallizes with two independent molecules in a triclinic cell. The cell parameters are: space group P1, a=10.100(2)Å, b=15.194(4) Å, c=19.948(5) Å, α=63.12(2)°, β=88.03(2)°, γ=88.61(2)°, Z=2, R=7.96% for 5140 data where |F_o|>3σ(F). The two independent molecules alternate in infinite columns formed by head-to-tail hydrogen bonding. The helices in the two independent molecules are quite similar to each other but one molecule is rotated ≈?123° about its helix axis with respect to the other. All the helical columns pack parallel to each other in the crystal. Replacement of the bulky Boc group does not lead to any major changes in conformation. Packing characteristics are also similar to those observed for similar helical peptides.     

Type II β-turn conformation of pivaloyl-L-prolyl-α-aminoisobutyryl-N-methylamide: Theoretical,spectroscopic, and X-ray studies

Pivaloyl-L -Pro-Aib-N-methylamide has been shown to possess one intramolecular hydrogen bond in (CD₃)₂SO solution, by ¹H-nmr methods, suggesting the existence of β-turns, with Pro-Aib as the corner residues. Theoretical conformational analysis suggests that Type II β-turn conformations are about 2 kcal mol^?1 more stable than Type III structures. A crystallographic study has established the Type II β-turn in the solid state. The molecule crystallizes in the space group P2₁ with a = 5.865 Å, b = 11.421 Å, c = 12.966 Å, β = 97.55°, and Z = 2. The structure has been refined to a final R value of 0.061. The Type II β-turn conformation is stabilized by an intramolecular 4 → 1 hydrogen bond between the methylamide NH and the pivaloyl CO group. The conformational angles are ?_Pro = ?57.8°, ψ_Pro = 139.3°, ?_Aib = 61.4°, and ψ_Aib = 25.1°. The Type II β-turn conformation for Pro-Aib in this peptide is compared with the Type III structures observed for the same segment in larger peptides.     

Molecular dynamics of sickle and normal hemoglobins

Molecular dynamics (MD) simulations have been carried out for 62.5 ps on crystal structures of deoxy sickle cell hemoglobin (HbS) and normal deoxy hemoglobin (HbA) using the CHARMM MD algorithm, with a time step of 0.001 ps. In the trajectory analysis of the 12.5–62.5 (50 ps) simulation, oscillations of the radius of gyration and solvent-accessible surface area were calculated. HbS exhibited a general contraction during the simulation, while HbA exhibited a nearly constant size. The average deviations of simulated structures from the starting structures were found to be 1.8 Å for HbA and 2.3 Å for HbS. The average rms amplitudes of atomic motions (atomic flexibility) were about 0.7 Å for HbA and about 1.0 Å for HbS. The amplitudes of backbone motion correlate well with temperature factors derived from x-ray crystallography. A comparison of flexibility between the α- and β-chains in both HbA and HbS indicates that the β-chains generally exhibited greater flexibility than the α-chains, and that the HbS β-chains exhibit greater flexibility in the N-terminal and D- and F-helix regions than do those of HbA. The average amplitude of backbone torsional oscillations was about 9°, a value comparable with that of other simulations, with enhanced torsional oscillation occurring primarily at the ends of helices or in loop regions between helices. Comparison of atomic flexibility and torsional oscillation results suggests that the increased β-chain flexibility results from relatively concerted motions of secondary structure elements. The increased flexibility may play an important role in HbS polymerization. Time course analysis of conformational energy of association, hydrogen bonding and hydrophobic bonding (as calculated from solvent accessibility) shows that all three of these factors contribute to the stability of subunit association for both hemoglobins. © 1993 John Wiley & Sons, Inc.     

Crystal and molecular structure of cyclohepta-amylose dodecahydrate

β-Cyclodextrin (cyclohepta-amylose, β-CD) is a torus-shaped, cyclic heptasaccharide consisting of (1→4)-linked α-d-glucopyranosyl residues. It is able to form inclusion complexes with small molecules in aqueous solution because of its annular aperture (width, 6.2 Å). β-Cyclodextrin dodecahydrate, the “empty” β-CD, crystallises from water in space group P2₁, with cell constants a = 21.29(2), b = 10.33(1), c = 15.10(2) Å, and β = 112.3(5)°. A total of 5189 X-ray counter-data were collected on a four-circle diffractometer. The crystal structure was solved on the basis of the highly isomorphous β-CD · 2HI · 8H₂O adduct, and the atomic parameters were refined by the full matrix, least-squares method to R = 7.3% for all data. The crystal structure belongs to the cage type. The β-CD macrocycle exists in an open, circular conformation stabilised by intramolecular hydrogen-bonds between HO-2 and HO-3 of adjacent glucosyl residues; four of the seven HO-6 groups are in the favoured (?)gauche orientation with respect to O-5, two are in the (+)gauche orientation, and one is disordered over these two orientations. The 6.5 water molecules within the cavity are distributed over 8 sites and display extensive thermal motion which is probably correlated with statistical disorder.     

The Conformation of Single Stranded Oligonucleotides and of Oligonucleotide-Oligopeptide Complexes from their Rotation Relaxation in the Nanosecond Time Range

Abstract

The conformation of single stranded oligonucleotides is analysed by measurements of their rotation time constants. The oligomers are aligned to some degree by short electric field pulses; after pulse termination the transition to a random orientation is followed by measurements of the linear dichroism. An efficient deconvolution procedure is developed for evaluation of the experimental data obtained in the ns-time range. The increase of rotation time constants observed for chain lengths in the range from 14 to 22 residues are interpreted according to a weakly bending rod model providing a persistence length and a Stokes' diameter. The Stokes' diameters obtained for ribo- and deoxyriboadenylates are about 13Å, in approximate agreement with the expectation for a single stranded helix. The persistence length L = 53Å corresponding to ～16 nucleotide residues found for riboadenylates at 2°C appears to reflect relatively strong stacking interactions at this temperature. However, a comparison with the average length of stacked residues evaluated from available thermodynamic parameters of base stacking indicate that unstacked residues are not completely flexible. Apparently the ribose-phosphate chain provides an essential contribution to the stiffness of oligomers and polymers, even when the bases are unstacked. Addition of 100μM Mg²⁺ leads to an increase of the persistence length to 88Å. Corresponding measurements with deoxyriboadenylates show a slightly lower value of the persistence length than that found for riboadenylates. Addition of LysTrpLys and LysTyrLys to A(pA)₁₉ leads to an increase of the rotation time constant, which corresponds approximately to a length increment by one residue per bound peptide. Since controls performed with LysLeuLys do not show any similar effect, the increase of the time constants induced by LysTrpLys and LysTyrLys is attributed to intercalation of the aromatic amino acids.     

Structures of the “open” and “closed” state of trypanosomal triosephosphate isomerase,as observed in a new crystal form: Implications for the reaction mechanism

The structure of trypanosomal triosephosphate isomerase (TIM)has been solved at a resolution of 2.1Å in a new crystal form grown at pH 8.8 from PEG6000. In this new crystal form (space group C2, cell dimensions 94.8 Å, 48.3 Å, 131.0 Å, 90.0°, 100.3°, 90.0°), TIM is present in a ligand-free state. The asymmetric unit consists of two TIM subunits. Each of these subunits is part of a dimer which is sitting on a crystallographic twofold axis, such that the crystal packing is formed from two TIM dimers in two distinct environments. The two constituent monomers of a given dimer are, therefore, crystallographically equivalent. In the ligand-free state of TIM in this crystal form, the two types of dimer are very similar in structure, with the flexible loops in the “Open” conformation. For one dimer (termed molecule-1), the flexible loop (loop-6) is involved in crystal contacts. Crystals of this type have been used in soaking experiments with 0.4 M ammonium sulphate (studied at 2.4 Å resolution), and with 40 μM phosphoglycolohydroxamate (studied at 2.5 Å resolution). It is found that transfer to 0.4 M ammonuum sulphate (equal to 80 times the Ki of sulphate for TIM), gives rise to significant sulphate binding at the active site of one dimer (termed molecule-2), and less significant binding at the active site of the other. In neither dimer does sulphate induce a “closed” conformation. In a mother liquor containing 40 μM phosphoglycolohydroxamate (equal to 10 times the Ki of phosphoglycolohydroxamate for TIM), an inhibitor molecule binds at the active site of only that dimer of which the flexible loop is free from crystal contacts (molecule-2). In this dimer, it induces a closed conformation. These three structures are compared and discussed with respect to the mode of binding of ligand in the active site as well as with respect to the conformational changes resulting from ligand binding. © 1993 Wiley-Liss, Inc.     

Flow-induced conformational changes and phase behavior of aqueous poly-L-lysine solutions

Poly-L -lysine exists as an α-helix at high pH and a random coil at neutral pH. When the α-helix is heated above 27°C, the macromolecule undergoes a conformational transition to a β-sheet. In this study, the stability of the secondary structure of poly-L -lysine in solutions subjected to shear flow, at temperatures below the α-helix to β-sheet transition temperature, were examined using Raman spectroscopy and CD. Solutions initially in the α-helical state showed time-dependent increases in viscosity with shearing, rising as much as an order of magnitude. Visual observation and turbidity measurements showed the formation of a gel-like phase under flow. Laser Raman measurements demonstrated the presence of small amounts of β-sheet structure evidenced by the amide I band at 1666 cm⁻¹. CD measurements indicated that solutions of predominantly α-helical conformation at 20°C transformed into 85% α-helix and 15% β-sheet after being sheared for 20 min. However, on continued shearing the content of β-sheet conformation decreased. The observed phenomena were explained in terms of a “zipping-up” molecular model based on flow enhanced hydrophobic interactions similar to that observed in gel-forming flexible polymers. © 1998 John Wiley & Sons, Inc. Biopoly 45: 239–246, 1998     

Identification and mass analysis of human fibrinogen molecules and their domains by scanning transmission electron microscopy

The domainal substructure and molecular conformation of human fibrinogen have been investigated by evaluating scanning transmission electron microscopic images of freeze-dried or negatively contrasted native fibrinogen (fractions I-4 and I-9), glutaraldehyde-treated fibrinogen, or plasmic core fragments D₁ and E₂. Although some unstained freeze-dried native or glutaraldehyde-treated fibrinogen molecules were relatively compact and even occasionally spheroidal, typical images were elongated symmetrical tridomainal structures 460 Å ± 20 Å in length; frequently they were bent into a variety of elongated though non-linear arrangements. Their identification as monomolecular forms of fibrinogen by scanning transmission electron microscopic mass measurements resolved uncertainties relating to the identity of such objects as single molecules. The central domains of fraction I-4 molecules had a greater mass than those of fraction I-9 (1.01 × 10⁵M_rversus 7.5 × 10 M_r, respectively). This difference accounted for the observed mass difference between fraction I-4 and fraction I-9 molecules (i.e. 3.27 × 10⁵M_rversus 2.97 × 10⁵M_r, respectively) and suggested that the COOH-terminal region of the Aα chain (major portions of which are always absent from fraction I-9 molecules) is situated within the mass integration radius for the central domain. When the COOH-terminal region of the Aα chain was present it appeared in negative stain as a thread-like structure originating between the middle and outer domains and extending toward the central domain, sometimes appearing to wind around the long axis.The outer domains of negatively stained molecules resembled negatively stained images of fragment D₁ and could frequently be resolved into at least two discrete subdomains, forming an oblong structure usually canted at an angle of ～120 ° to 150 ° relative to the long axis. Our findings are consistent with prevailing tridomainal structural models of fibrinogen and suggest that these molecules are flexible and may exist in unfolded configurations, or as relatively compact, partially or completely folded forms.     

An X-ray diffraction study of poly(L-lysine hydrobromide)

A structural study of the synthetic polypeptide poly(L -lysine hydrobromide) has been made by X-ray fiber techniques. The investigation was undertaken to determine whelther this polymer undergoes conformational transitions as a function of hydration in a manner similar to other chemically related basic polypeptides. Specifically, a comparison with the previously reported structures of the hydrochloride form of poly(L -lysine) was sought. Homogeneous powder mixtures with various amounts of water and oriented fibers of poly(L -lysine hydrobromide) at different relative humidities were X-ray photographed. Reversible transitions amorphous state ? β-pleated sheet ? α-helix ? isotropic solution as a function of increasing/decreasing degrees of hydration were found. The β-pleated-sheet conformation was observed between 33% and 76% relative humidities (containing about one and three molecules of water per residue, respectively). Each pleated sheet was formed by “antiparallel” chains, and the sheets were piled up along the b-axis. The spacings of this conformation did not vary appreciably with hydration. The observed reflections at 52% relative humidity (1.4 molecules of water per residue) could be indexed satisfactorily in terms of an orthorhombic unit cell, of space group P2₁22₁, with a = 9.52 Å, b = 16.44 Å, and c = 6.80 Å. These dimensions were shown by models to be compatible with the proposed structure. The α-helix conformation was present in specimens photographed at 76% relative humidity and up, and containing between three and fifteen molecules of water per residue. The helices were packed parallel to each other in a hexagonal array but randomly along or about their lengths. Increasing the hydration from five to fifteen molecules of water per residue causes the a-axis to increase from 16.9 to 20.8 Å. Twenty molecules of water per residue produced an isotropic solution. Despite some structural differences between the hydrobromide and hydrochloride forms it is concluded that the role played by the anions is mainly related to determining the water content levels at which conformational changes occur. Therefore, the anions do not significantly influence the prevailing conformation in this particular system, but might affect the packing arrangement of the polypeptide chains.     

Theory of H1-mediated control of higher orders of structure in chromatin

It is known that the lysine-rich histone H1 induces both higher orders of folding in chromatin and donut shapes in DNA. However, these phenomena occur only on the high-salt side of a narrow transition range located at about 0.02M salt. Previous theoretical analyses of the ionic-strength dependencies of DNA persistence length and denaturation rate have provided the information that the basic rigid-rod unit in high-molecular-weight DNA is a segment 60 base pairs in length and that if the phosphate charge is neutralized, this segment will spontaneously adopt a bent conformation with radius of curvature 170 Å. On the assumption that an H1 molecule does not completely neutralize the DNA charge in its vicinity, the theory has been extended here to determine the onset of spontaneous bending as a function of salt concentration and extent of phosphate neutralization. A salt transition of the kind observed has been found for the realistic value of 82% charge neutralization, with the actual value likely to be in the neighborhood of 90%, as suggested by the measurements of Wilson and Bloomfield.¹ It is recalled that the spacer DNA length in chromatin is of about the same length as the DNA rigid-rod unit. If binding of H1 to the spacer induces, as predicted, a bent conformation of radius about 170 Å, then the observed value of about 150 Å for the outer radius of the solenoid presently thought to be the basic mode of folding for a nucleosome chain can be understood as a reflection of the inherent maximum curvature of DNA in aqueous salt solution.     

Crystallization,crystal structure analysis and molecular model of the third domain of Japanese quail ovomucoid,a Kazal type inhibitor

The third domain of Japanese quail ovomucoid, a Kazal type inhibitor, has been crystallized and its crystal structure determined at 2.5 Å resolution using multiple isomorphous replacement techniques. The asymmetric unit contains four molecules. In the crystal the molecules are arranged in two slightly different octamers with approximate D4 symmetry. The molecules are held together mainly by interactions of the N-terminal residues, which form a novel secondary structural element, a β-channel.The molecule is globular with approximate dimensions 35 Å × 27 Å × 19 Å. The secondary structural elements are a double-stranded anti-parallel β-sheet of residues Pro22 to Gly32 and an α-helix from Asn33 to Ser44. The reactive site Lys18-Asp19 is located in an exposed loop. It is close to Asn33 at the N terminus of the helical segment. The polypeptide chain folding of ovomucoid bears some resemblance to other inhibitors in the existence of an anti-parallel double strand following the reactive site loop.