Helix-coil Transition Of Poly Research Articles

Crowding effect on helix-coil transition: Beyond entropic stabilization

We report circular dichroism measurements on the helix-coil transition of poly(L-glutamic acid) in solution with polyethylene glycol (PEG) as a crowding agent. The PEG solutions have been characterized by small angle neutron scattering and are well described by the picture of a network of mesh size ξ, usual for semi-dilute chains in good solvent. We show that the increase of PEG concentration stabilizes the helices and increases the transition temperature. But more unexpectedly, we also notice that the increase of concentration of crowding agent reduces the mean helix extent at the transition, or in other words reduces its cooperativity. This result cannot be taken into account for by an entropic stabilization mechanism. Comparing the mean length of helices at the transition and the mesh size of the PEG network, our results strongly suggest two regimes: helices shorter or longer than the mesh size.

Osmotically Induced Helix-Coil Transition in Poly(Glutamic Acid)

Protein folding and conformational changes are influenced by protein-water interactions and, as such, the energetics of protein function are necessarily linked to water activity. Here, we have chosen the helix-coil transition in poly(glutamic acid) as a model system to investigate the importance of hydration to protein structure by using the osmotic stress method combined with circular dichroism spectroscopy. Osmotic stress is applied using poly(ethylene glycol), molecular weight of 400, as the osmolyte. The energetics of the helix-coil transition under applied osmotic stress allows us to calculate the change in the number of preferentially included water molecules per residue accompanying the thermally induced conformational change. We find that osmotic stress raises the helix-coil transition temperature by favoring the more compact α-helical state over the more hydrated coil state. The contribution of other forces to α-helix stability also are explored by varying pH and studying a random copolymer, poly(glutamic acid- r-alanine). In this article, we clearly show the influence of osmotic pressure on the peptide folding equilibrium. Our results suggest that to study protein folding in vitro, the osmotic pressure, in addition to pH and salt concentration, should be controlled to better approximate the crowded environment inside cells.

Comparison of the Heat- and Pressure-Induced Helix−Coil Transition of Two DNA Copolymers

The helix-coil transition of poly[d(I-C)] and poly[d(A-T)] was studied as a function of hydrostatic pressure, temperature, and sodium ion concentration. These studies were undertaken in light of a recently published phase diagram for double stranded nucleic acids [Dubins et al. J. Am. Chem. Soc. 2001, 123, 9254-9259]. The sign and magnitude of the volume change for the heat-induced helix-coil transition, DeltaV(T), of poly[d(I-C)] and poly[d(A-T)] were dependent on the helix-coil transition temperature, T(M), at atmospheric pressure. The sign of DeltaV(T) changed from negative to positive as T(M) was increased by increasing the sodium ion concentration. For poly[d(I-C)], DeltaV(T) = 0 cm(3) mol(-1), when the sodium ion concentration is such that the spectroscopically monitored T(M) = 55 degrees C at atmospheric pressure. For poly[d(A-T)], the value of DeltaV(T) = 0 under conditions such that T(M) = 47 degrees C at atmospheric pressure. Negative values of DeltaV(T) imply that the helical form is destabilized at high pressure. Under experimental conditions where the DeltaV(T) for the transition is negative, the transition could be caused by increasing the pressure under isothermal conditions. At temperatures below T(M) measured at atmospheric pressure the midpoint of the pressure-induced helix-coil transition, P(M), decreases with increasing temperature. The volume change of the pressure-induced transitions helix-coil transition, DeltaV(P), was calculated assuming a two-state model. The magnitude of DeltaV(P) (per cooperative length) was much larger than the volume change (per base pair) measured for the heat-induced transition, DeltaV(T), calculated using the Clapeyron equation. The ratio of these two volume changes was used to calculate the cooperative length for the pressure-induced transition. This parameter depends strongly on temperature, becoming greater closer to T(M) measured at atmospheric pressure. At temperatures approaching T(M) the magnitude of the cooperative length of the pressure-induced transition is approximately twice that observed for the heat-induced transition (N(T)). On the basis of the temperature dependence of the DeltaV(T) for the two polymers the coefficient of thermal expansion of the two polymers was found to be 0.17 and 0.16 cm(3) K(-1) mol(-1) for poly[d(I-C)] and poly[d(A-T)], respectively.

Polarographic behavior of Cd 2+ and Zn 2+—PGA complexes. Polarographic and circular dichroism spectroscopic studies on the Cd 2+ and/or Zn 2+—poly(α-L-glutamic acid) complex in aqueous solution

The helix-coil transition of poly(α-L-glutamic acid) (PGA) induced by Cd 2+ and/or Zn 2+ was investigated by direct current (d.c.) polarography and circular dichroism (CD) spectroscopy as a function of pH and the mixing ratio of glutamic acid residue to Cd 2+ and/or Zn 2+. The diffusion current for Cd 2+ or Zn 2+ declined sharply in the pH range of c. 4–7 due to the formation of the metal ion-PGA helical complex. The helix contents of PGA, estimated by the decrease in polarographic current, are in good agreement with results obtained by circular dichroism in the neutral pH region. The degree of helicity induced by either of these two ions can also be determined independently by polarography. The size of the complex was calculated by the Ilkovič equation and depends strongly on the mixing ratio of glutamic acid to cation. In the neutral pH region, the relationship between the half-wave potentials and the concentration of PGA indicates that two peptide residues coordinate approximately to one Cd 2+.

Prediction of the thermodynamics of protein unfolding: the helix-coil transition of poly(L-alanine).

The method given earlier for predicting the thermodynamics of protein unfolding from the x-ray structure of a protein is applied here to the poly(L-alanine) helix. First, the fitting parameters derived earlier from a data base of 10 proteins were used to predict the unfolding thermodynamics of 4 other proteins. The agreement between the observed and predicted values is comparable to that found for the 10 proteins studied initially. Next, the temperature dependences of the Gibbs energy and enthalpy changes for unfolding of bacteriophage T4 lysozyme were predicted and compared with data in the literature. The predicted and observed temperature dependences are similar and the predicted results indicate that cold denaturation should be observed at low temperatures, as observed recently for a T4 lysozyme mutant. The fitting parameters derived from thermodynamic data for protein unfolding and for hydration of model compounds were used to predict the unfolding thermodynamics of the poly(L-alanine) helix. The results predict that helix formation is enthalpy-driven, and the predicted enthalpy change for unfolding (0.86 kcal per mol per residue) is close to the value found in a recent calorimetric study of a 50-residue alanine-rich helix.

Bimodal distribution of relaxational modes for the helix-coil transition in poly(oxymethylene)

The helix-coil transition in poly(oxymethy1ene) has been investigated by using the recently developed dynamic rotational isomeric state (DRIS) model and the related matrix multiplication technique. The conditional probability P,(h/h) that a chain originally in the perfect helical form will preserve its state unchanged at a later time T is evaluated for various chain lengths and temperatures. The transition to the coiled structure in an originally fully ordered helix is found to exhibit a bimodal relaxation, indicated by a plateau in the time decay curves of the conditional probabilities, before restoration of equilibrium distribution. This intermediate quasi-steady state preceding full relaxation is attributed to an equilibrium between perfect helices and chains possessing a single defect in the form of a trans bond, interrupting sequences of gauche bonds of the same sign.

Polarographic study of poly(α-D-glutamic acid) in acid solutions

A reduction wave with a half-wave potential of about −1.45V ( versus n.h.e.) was observed for the first time in tast polarographic measurement of poly(α-D-glutamic acid), poly(Glu), in acid solutions. This wave was diffusion-controlled and its height increased with increasing H + ion concentration. The reduction wave was examined in detail by carrying out polarographic measurements both for carboxyl-containing compounds such as malonic acid, EDTA, and poly-acrylic acid, and for poly(L-ornithine), a polypeptide which does not contain any carboxyl groups. Cyclic voltammetry was also performed for poly(Glu) in acid solutions. The present experiments strongly indicated that poly(Glu) forms a proton chelate complex in acid solution. This complex is responsible for the appearance of a polarographic reduction wave at about −1.45 V ( versus n.h.e.). The concept of intramolecular proton chelates was also supported by analysis of the limiting current with the aid of the Ilkovič equation, from which the diffusion coefficient of poly(Glu) was obtained. The correlation between the limiting current at −1.45 V and the helix-coil transition curve of poly(Glu) was examined. The reduction waves at −1.45 V appeared in the pH region where the helix-coil transition of poly(Glu) takes place. On the basis of these results, a possible chelated structure of poly(Glu) in acid solution is discussed.

The Visible-UV Absorption and Circular Dichroism Studies of Poly(α-l-glutamic acid)–Copper(II) Complexes with Emphasis on the Extrinsic Cotton Effect

Abstract The visible-UV absorption and circular dichroism, CD, of poly(α-l-glutamic acid)–Cu(II) solutions have been measured at 25 °C in the pH range 7 to 3.5 and at the mixing ratio, R, (Glu residues-to-Cu2+) 32 to 4. Complex formation between poly (Glu) and Cu2+ was evident from the three absorption bands of bound Cu(II) around 700, 370, and 250 nm. The molar absorption coefficients, ε, varied with pH and R in a complicated but systematic manner. The pH dependence of ε, however, was not in parallel with the helix-coil transition of poly(Glu)–Cu(II) complexes. The extrinsic CD bands of bound Cu(II) have been detected in the three absorption band regions, and found to depend on both pH and R. The optical dissymmetry factor indicated that the 700 and 250 nm absorption bands of bound Cu(II) are associated with multiple CD bands of opposite signs. The intrinsic CD showed that the Cu(II)- and pH-induced helices of the poly(Glu)–Cu(II) complex are alike regarding the polymer backbone.

Helix-coil stability constants for the naturally occurring amino acids in water. 16. Aspartic acid parameters from random poly(hydroxybutylglutamine-co-L-aspartic acid).

The synthesis and characterization of water-soluble random copolymers containing L-aspartic acid with N5-(4-hydroxybutyl)-l-glutamine, and the thermally induced helix-coil transitions of these copolymers in water and in 0.1 N KCl, are described. The incorporation of L-aspartic acid was found to decrease the helix content of the polymer at both high and low pH, in water and also in 0.1 N KCl. The Zimm-Bragg parameters sigma and s for the helix-coil transition in poly(L-aspartic acid) in water and in 0.1 N KCl were deduced from an analysis of the melting curves of the copolymers in the manner described in earlier papers. Corrections were made for the presence of a small amount of racemized aspartic acid, using data from random copolymers containing D-aspartic acid as the guest residue. The computed values of s indicate that L-aspartic acid destabilizes helical sequences at all temperatures in the range of 0-70 degrees C. Titrations of the copolymers and of N-acetyl-N'-methyl-L-aspartic acid amide in 0.1 N KCl are described.

Helix-coil stability constants for the naturally occurring amino acids in water. 15 Arginine parameters from random poly(hydroxybutylglutamine-co-L-arginine).

Water-soluble copolymers containing L-arginine and N5-(4-hydroxybutyl)-L-glutamine were prepared by copolymerization of the N-carboxy-alpha-amino acid anhydrides of Ndelta-tert-butyloxycarbonyl-L-ornithine and gamma-benzyl L-glutamate, followed by aminolysis with 4-amino-1-butanol, by removal of the tert-butyloxycarbonyl protecting group, and by treatment with O-methylisourea. The copolymers were fractionated and characterized, and the thermally induced helix-coil transitions of these copolymers were studied in water at neutral pH in the presence and in the absence of KCl. The Zimm-Bragg parameters sigma and s for the helix-coil transition in poly(L-arginine) in aqueous solution were deduced from an analysis of the melting curves of the copolymers in the manner described in earlier papers. The computed values of s indicate that L-arginine is a weak helix-making residue at low temperature and a weak helix-breaking residue at high temperature in aqueous solution. The results were found to be in good agreement with those obtained earlier in conformational analyses of arginyl residues in proteins.

Helix-coil stability constants for the naturally occurring amino acids in water. 11. Lysine parameters from random poly(hydroxybutylglutamine-co-L-lysine).

The synthesis and characterization of water-soluble random copolymers containing L-lysine with N5-(4-hydroxybutyl)-L-glutamine, and the thermally induced helix-coil transitions of these copolymers in water, are described. The incorporation of L-lysine was found to decrease the helix content of the polymers at neutral pH. The Zimm-Bragg parameters sigma and s for the helix-coil transition in poly(L-lysine) in water were deduced from an analysis of the melting curves in the manner described in earlier papers. The computed values of s indicate that, in the temperature range of 0-60 degrees C, lysine has a tendency to destabilize helical sequences, this tendency being minimal at approximately 25 degrees C and increasing at lower and higher temperatures.

Thermodynamic and kinetic parameters of ion condensation to polynucleotides

The coupling of ion binding to the single strand helix-coil transition in poly (A) and poly (C) is used to obtain information about both processes by ion titration and field-jump relaxation methods. Characterisation of the field-jump relaxation in poly (C) at various concentrations of monovalent ions leads to the evaluation of a stability constant K = 71 M(-1) for the ion binding to the polymer. the rate constant of helix formation is found to be 1.3 X 10(7)s(-1), whereas the dissociation rate is 1.0 X 10(6)s(-1). Similar data are preseented for poly (A) and poly (dA). The interaction of Mg(++) and Ca(++) with poly (A) and poly (C) is measured by a titration method using the polymer absorbance for the indication of binding. The data can be represented by a model with independent binding "sites". The stability constants increase with decreasing salt concentration from 2.7 X 10(4) M(-1) at medium ionic strengths up to 2.7 X 10(7) M(-1) at low ionic strength. The number of ions bound per nucleotide residue is in the range 0.2 to 0.3. Relaxation time constants associated with Mg(++) binding are characterised over a broad range of Mg(++) concentrations form 5 muM to 500 muM. The observed concentration dependence supports the conclusion on the number of binding places inferred from equilibrium titrations. The rate of Mg(++) and Ca(++) association to the polymer is close to the limit of diffusion control kR =1 X 10(10) to 2 X 10(10) M(-1) s(-1)). This high rate demonstrates that Mg(++) and Ca(++) ions do not form inner-sphere complexes with the polynucleotides. Apparently the distance between two adjacent phosphates is too large for a simultaneous site binding of Mg(++) or Ca(++), and inner sphere complexation at a single phosphate seems to be too weak. The data support the view that the ions like Mg(++) and Ca(++) surround the polynucleotides in the form of a mobile ion cloud without site binding.

Molecular Theory of the Helix-Coil Transition in Poly(amino acids). IV. Evaluation and Analysis of s for Poly(L-valine) in the Absence and Presence of Water

Molecular Theory of the Helix-Coil Transition in Poly(amino acids). IV. Evaluation and Analysis of s for Poly(L-valine) in the Absence and Presence of Water

Carbon-13 nuclear magnetic resonance studies of polyamino acids: The helix-coil transition of poly- l-lysine

The helix-coil transition of poly- l-lysine hydrochloride ((Lys) n) in aqueous solution has been studied by 13C Fourier-transform nuclear magnetic resonance spectroscopy. As reference compounds dodeca- l-lysine hydrobromide ((Lys) − 12, tri- l-lysine hydrochloride ((Lys) 3), and l-lysine hydrochloride (Lys), have been also studied by the same method. It is found that 13C spin-lattice relaxation times t 1 of the carbonyl and the side-chain carbons decrease sharply at p D 10.2 which is the midpoint of the transition from the random-coil to the α-helix. Similarly the T 1 values of the carbonyl groups of (Lys) − 12 decrease at this point in a more moderate way, while no change is observed for those of the side-chain carbons. This is interpreted in terms of the reduced α-helicity involved for (Lys) − 12. The variation of 13C chemical shifts with p D for (Lys) n and (Lys) − 12 show the same trend:downfield shifts at higher p D. Furthermore, nonterminal and C-terminal residues of (Lys) 3 show similar behavior. Thus it is concluded that the 13C chemical shift changes are caused mainly by the p D changes and not by the conformational transition. Conversion from α-helix to β-structure by elevation of temperature at p D 11.2 results in narrowing and downfield shifts of the 13C resonances of (Lys) n .

Helix-Coil Transition of Poly(N ε-carbobenzoxy-L-lysine) in Solutions Containing Trifluoroacetic Acid

Helix-Coil Transition of Poly(N ^ε-carbobenzoxy-L-lysine) in Solutions Containing Trifluoroacetic Acid

Helix to coil transition in poly(γ-benzyl-L-glutamate)

The helix-coil transition of poly(γ-benzyl-L-glutamate), Mv=530.000, in mixture of chloroform (CHCl3) and dichloroacetic acid (DCA) was studied by measuring refractive index increments (dn/dc) and reduced sedimentation coefficients ([S]) as functions of the solvent composition.

The properties of the helix and coil forms of polyribouridylic acid and its halogenated analogs

1. 1. The helix-coil transition of poly U has been studied by various optical and hydrodynamic methods. It is concluded that the helix form of poly U is essentially that of a single strand folded back on itself — a hairpin. 2. 2. Poly U forms temperature labile complexes with various halogenated poly uridylic acids.

Helix-coil transition of poly- l-phenylalanine studied by nuclear magnetic resonance

This chapter discusses the application of nuclear magnetic resonance (NMR) to protein chemistry primarily in terms of the information that the technique can contribute to the protein structure and function. The chapter examines information about proteins that can be derived from NMR spectra, the premises and procedures necessary to interpret the observations. It also discusses whether NMR is merely another method for confirming conclusions reached by other methods or does it, in fact, permit findings not obtainable otherwise. With the success of selective deuteration of proteins, the two major problems of protein NMR spectroscopy—resolution and assignment of resonance lines—have been shown to be soluble at least in principle. It is to be expected that future difficulties in overcoming these problems will be proportional to the complexity of the protein. Success, however, is to be sought in experimental ingenuity rather than through further advances in instrumentation.