Thermodynamic parameters of helix-coil transition in polypeptide chains. I. Poly-(L-glutamic acid)

The helix-coil transitions for poly(L -glutamic acid) (PGA) in 0.2M NaCl and in its mixture with dioxane were studied by the methods of spectropolarimetry, viscometry, and potentiometric titration at different temperatures from 8 to 50°C. The enthalpy and entropy differences between the helical and coillike states of uncharged PGA molecules were determined from the curves of potentiometric titration. The temperature dependence of the cooperativity parameter σ was determined by two methods: from the sharpness of transition and from the dependence of the intrinsic viscosity on the helical content in the transition region. In 0.2MNaCl, σ= (2.5 ± 0.5) × 10^?3 and practically does not depend on temperature, i.e., the cooperativity of the helix-coil transition is connected mainly with the entropy decrease in initiating helical regions (ΔS_i ≈ ?12 is mole of helical regions). On the contrary, initiation of a helical region in the water-organic solvent mixture is accompanied by a considerable enthalpy increase.     

Thermodynamic parameters of helix-coil transition in polypeptide chains. II. Poly-L-lysine

The helix–coil transitions for poly-L -lysine (PL) were investigated by the methods of spectropolarimetry, viscometry and potentiometric titration in 0.2M NaCl at different temperatures as well as in 0.2MNaBr, 1MKCl, and in mixtures of 0.2MNaCl or NaBr with methanol at room temperature. The enthalpy and entropy differences between the helical and coillike states of uncharged PL molecules in 0.2.M NaCl were determined from the potentiometric titration curves. The cooperativity parameters σ for PL in different solvents were determined by two methods (from the sharpness of the transition and from the dependence of the intrinsic viscosity on the helical content in the transition region). In 0.2MNaCl σ has a value of (2.3 ± 0.5) × 10^?4 and does not depend on temperature, i.e., the cooperativity of the helix-coil transition, as for PGA, is mainly of an entropy origin (the initiating of the helical region is accompanied by the entropy decrease ΔS_i = ?12 eu/mole of helical regions). A comparison of the obtained results for PGA and PL with the molecular theories of the helix-coil transitions shows that the role of dipole-dipole interactions of nonneighboring peptide groups is greatly overestimated in these theories, leading to a considerable enthalpy contribution to the free energy of initiating helical regions which is not observed in the experiment.     

Potentiometric titrations and the helix-coil transition of poly(L-glutamic acid) and poly-L-lysine in aqueous salt solutions

The objective have been to establish if those ions which are known to change the stability of the structure of proteins, have any influence on the properties of ionizable polypeptides. Potentiometric titrations and complementary optical rotation data are presented for aqueous solutions of poly-L -lysine (PLL) in the presence of KSCN, KCl, and KF, and for poly(L -glutamic acid) (PLGA) in the presence of KSCN, KCl, and LiCl. The following measured quantities which are affected by salt concentration were obtained: intrinsic pK (pK₀), slope of pK_app versus degree of ionization (α) curves, the degree of ionization at which the helix to coil transition occurs, and the free energy of this transition for the uncharged molecule (δG°_hel). The effects of nonspecific salts (KCl and LiCl for PLL and KSCN and KCl for PLGA) are small, and about, as expected from general electrostatic considerations. In line with the observations made with isoelectric and cat ionic collagen, specific, effects were noted with KSCN–PLL and with LiCl–PLGA. In the presence of KSCN, the poly-L -lysine helix becomes stabilized at much lower degree of ionization than in the presence of KCl, and the slope of the pK_app versus α plots is greatly reduced. However, ΔG°_hel (for the uncharged molecule) is not affected, and pK₀ is only slightly higher. We interpret these data in terms of binding of SCN^? primarily to the side-chain amino groups (both to R? NH₃⁺ and to R? NH₂) solutions. (L -glutamic acid) in LiCl solution has its transition at the same α value as in KCl solution. However, both the slopes of the pK_app versus α plots and the absolute values of ΔG°_hel are lower than in KCl solution. We interpret these results in terms of binding of Li⁺ to side chains as well as to the peptide bond.     

Thermodynamic parameters for the helix-coil thermal transition of ribonuclease-S-peptide and derivatives from 1H-NMR data

Values for the thermodynamic quantities, ΔH° = 11.8 ± 2.0 Kcal/mole and ΔS° = 43.6 ± 6.0 e.u., of the 3-13 helix–coil equilibrium of isolated S-peptide (19 residue N-terminal fragment of ribonuclease A) in aqueous solution (3 m M, 1M NaCl, pD 5.4) have been determined from a joint analysis of the Thr 3γ, Ala 6β, Phe 8meta, and Phe 8para ¹H chemical shift vs temperature curves (?7 to 80°C) in several aqueous–trifluorethanol mixtures. Chemical shifts in the coil and in the helix have been determined for up to 16 protons belonging to the 3-13 fragment. Thermodynamic parameters have also been determined for C-peptide (13 residue fragment) and a number of S-peptide derivatives. From the variation of the values of the thermodynamic parameters at pD 2.5, 5.4, and 8.0, a quantitation of the two helix-stabilizing side-chain interactions can be made: (1) Δ(ΔH°) ? 5 Kcal/mole and Δ(ΔS°) ? 18 e.u. for the salt bridge Glu 2^? … Arg 10⁺ and (2) Δ(ΔH°) ? 3 Kcal/mole and Δ(ΔS°) = 9 e.u. for the one in which the His 12⁺ imidazolium group is involved, presumably a partial stacking with the Phe 8 side chain.     

Thermal and charge-induced coil to -helix transition of poly-L-glutamic acid and random L-glutamic acid-L-alanine copolymers

Thermal and charge induced random coil to α-helix transitions of poly-L -glutamic acid (PGA) were measured by optical rotatory dispersion in various solvents. The data of PGA in 0.1M Nacl were analyzed by the Zimm-Rice theory. Enthalpy and entropy changes for the coil-to-helix transition in the unionized state were obtained: ΔH° = ?1020 ± 100 cal/residue mole; ΔH° = ?3.0 ± 0.4 e.u./residue mole. The initiation parameter, σ, of the Zimm-Rice theory was given by a value of 5 ± 1 × 10^-3. Random copolymers of L -glutamic acid and L -alanine containing 10, 30, and 40 molar percents of alanyl residue were synthesized. Stabilities of α-helix of the copolymers were compared to that of PGA. In water and water-ethanol solutions, stabilities of the polymers were almost equal after the simple correction about the ionized charge density of the polymers. In 0.1 M NaCl solution these copolymers showed some deviations from the transition curve of PGA, which would suggest the hydrophobic contribution of the alanyl residues.     

Solution properties of synthetic polypeptides. XII. Enthalpy changes accompanying helix-coil transition of polypeptide

For helix-coil transitions of polypeptide in binary mixtures consisting of helix-forming solvent and coil solvent, the transition enthalpy ΔH(T,x) has been found to depend significantly on temperature (T) and solvent composition (x). For such systems, calorimetric measurements may yield some averages of ΔH(T,x) which are no longer amenable to direct comparison with ΔH itself. Theoretical equations relating calorimetric data to ΔH(T,x) are derived and tested favorably with experimental data. It is demonstrated that the transition enthaply from heat capacity measurements is approximately equal to ΔH_cfm, while those from heat of dilution and heat of solution measurements are equal to ΔH_c. Here ΔH_c denotes the value of ΔH at the transition point and f_m represents the maximum helical content attained in a thermally induced transition. The discrepancies among calorimetric data are also discussed.     

Conformational properties of L-leucine,L-isoleucine,and L-norleucine side chains in L-lysine copolymers

The structure inducing properties of L -leucine, L -isoleucine, and L -norleucine residues incorporated into poly(L -lysine) were investigated by the observation of the circular dichroism of the respective random copolypeptides. The comparison involves the coil-helix transition in water/methanol mixtures, the formation of ordered structures at higher pH, and the kinetics of the α-helix to β-conformation transition of the leucine and norleucine copolymers induced by temperature changes at pH 10.5. The results confirm the known properties of the leucine residue, strongly supporting the α-helix conformation. They also support the idea that the isoleucine residue is one of the most powerful candidates for β-structure formation, and they show that the unbranched norleucine residue has intermediate properties. The results are discussed on the basis of steric and hydrophobic properties of the three side chains.     

Thermodynamic values of the helix-coil transition of DNA in the presence of quaternary ammonium salt

The knowledge of the enthalpy and entropy of the helix-coil transition in DNA is necessary for the understanding of the stabilization of its native conformation in solution. Reported here is the transition temperature Tm, the transition enthalpy deltaH, determined with the help of an adiabatic scanning calorimeter, the transition entropy deltaS and the breadth of the helix-coil transition as a function of tetramethyl and tetraethyl ammonium chloride concentration.     

Comparison of the helix-coil transition of a titrating polypeptide in aqueous solutions and at the air-water interface

The transition from alpha-helix to random coil of the titrating polyamino acid co-poly-L-(lysine, phenylalanine), (p-(Lys,Phe)), has been investigated as a function of pH and ionic strength in aqueous solution and at the air-water interface by means of circular dichroism (CD) spectroscopy and the Langmuir surface film balance technique. The results strongly suggest that the helix-coil transition for peptides at the air-water interface can be determined by using the two-dimensional Flory exponent, nu, to express the pH dependent peptide surface conformation. The helix-coil titration curve of p-(Lys,Phe) shifts approximately 2.5 pH units towards lower pH at the air-water interface, as compared with the bulk solution. This finding is of relevance for the understanding of conformation and conformational changes of membrane-transporting and membrane penetrating peptides as well as for the use of peptides in molecular devices.     

Pressure dependence of the helix-coil transition temperature for polynucleic acid helices

Thermal denaturation of DNA's and the corresponding helix–coil transformation of artificial polyribonucleic and polydeoxyribonucleic acids have been studied extensively both theoretically^1–13 and experimentally. ^14–30 Much less work has been carried out on the properties of these polynucleic acids at high pressure, and in particular, on the presure dependence of the helix–coil transition temperature.^31–33 Light-scattering techniques have been used in this study to measure the pressure dependence of the helix–coil transition temperature of the two- and three-stranded helices of polyriboadenylic and polyribouridilic acids and of calf thymus DNA. From the slopes of the transition temperature vs. pressure curves and heats of transition obtained from the literature,^20,34 the following volume changes from these helix–coil transitions have been obtained: (a) ?0.96 cc/mole of nucleotide base pairs for the poly (A + U) transition, (b) +0.35 cc/mole of nucleotide base trios for the poly (A + 2U) transition, and (c) +2.7 cc/mole of nucleotide base pairs for the DNA transition. The relative magnitudes and signs of these volume changes which show that poly (A + U) is destabilized by increased pressure, whereas poly (A + 2U) and calf thymus DNA are stabilized by increased pressure, indicates that further development of the helix–coil transition theory for polynucleotides is needed.     

Relaxation kinetics of the triple-stranded helix-coil transition at short chain length. A model including the staggering of chains.

The relaxation kinetics of a staggering zipper model are presented, in which the formation of helical nuclei is regarded to be rate-limiting and in which the sliding of strands along the helices is prohibited. Instead a realignment of staggered chains is only possible via complete uncoiling. While maintaining the third order of the initial reaction as in an all-or-none mechanism, the model predicts a large range of relaxation times, which contribute to the mean relaxation time according to the stability of the individual species and which are weakly coupled in the range of small amounts of helical content. The model can easily be compared to experimental results and agrees well with relaxation data obtained from a triple-helical peptide fragment of collagen. It may be readily expanded to other multistranded helix ? coil transitions with steady-state formation of the individual species and it suggests that the fraying of the helix ends is hidden by the fast-relaxation times due to the equilibration of the shortest helical species.     

Helix-coil transition of Poly(L-glutamic acid) in N-methylacetamide

The folding of randomly coiled poly(L -glutamic acid) to the helical state has been studied in N-methylacetamide by titration methods. Since this solvent would be expected to form amide-peptide group hydrogen bonds with the unfolded form of the polymer, to a first approximation no helix stabilization could come from intrapolymer hydrogen bonds. The titration data, collected from 30 to 70°C yield the following values per residue for the thermodynamic parameters governing the coil-helix reaction for the uncharged polymer: ΔG_30°C°, ?1. 9 ± 0.1 kcal; Δ H°, 0 ± 0.1 kcal; ΔS_30°C°, 6.3 ± 0.6 eu. In N-methyl acetamide, the helix is an order of magnitude more stable than in water, and this stabilization appears to be entirely the result of the entropy gained by solvent molecules which are released from the polymer upon folding.     

Kinetics of the helix-coil transition of a polypeptide with non-ionic side groups, derived from ultrasonic relaxation measurements.

Ultrasonic absorption and velocity dispersion curves have been measured in the temperature induced helix-coil transition range of poly-N5-(3-hydroxypropyl)-L-glutamine in a methanol/water mixture. The results clearly reflect an effect due to the kinetics of the conformational conversion. A practically single relaxation time is observed which passes through a maximum when plotted versus the degree of transition. This maximum occurs at definitely less than 50% helix as predicted for by the theory for the comparatively short chain length involved here. The results are discussed in relation to previous theoretical and experimental findings.     

Thermodynamic parameters of complexation of lanthanoid(III) with ascorbic acid

The thermodynamic parameters of formation of the 1:1 complex between trivalent lanthanide cations and ascorbic acid have been determined for 0.1 M and 2.0 M ionic strengths at 25 °C by potentiometric and calorimetric titration. Comparison of these values with data for other organic ligands indicates that in the complex LnAsc²⁺ the ascorbate functions as an inner sphere monodentate ligand more similar to benzoate than to kojate.     

Synthesis and characterization of biodegradable amphiphilic triblock copolymers containing L-glutamic acid units

A novel biodegradable amphiphilic triblock copolymer bearing pendant carboxyl groups PLGG-PEG-PLGG was successfully prepared by ring-opening copolymerization of l-lactide (LA) with (3s)-benzoxylcarbonylethyl-morpholine-2, 5-dione (BEMD) in the presence of dihydroxyl poly(ethylene glycol) (PEG) as a macroinitiator in bulk at 130 degrees C using SnOct(2) as catalyst and by subsequent catalytic hydrogenation. The copolymer could form micelles in aqueous solution with the cmc dependent on the composition of the copolymer. The micelles exhibited a homogeneous spherical morphology and a unimodal size distribution. Their degradation rate in the presence of proteinase K was faster than that of PLA, and they showed a low degree of cytotoxicity to the articular cartilage cells. This biodegradable amphiphilic block copolymer with pendant carboxyl groups is capable of further modification and is expected to facilitate a variety of potential biomedical applications, such as drug carriers, tissue engineering, etc.