Proton magnetic relaxation in solid poly(L-alanine), poly(L-leucine), poly(L-valine), and polyglycine

Molecular motion in solid poly(L -alanine), Poly(L -leucine), poly(L -valine), and polyglycine has been investigated through measurement of the portion spin-lattice relaxation time at 30 and 60 MHz between 110 and 350°K. Rapid random reoriention of sied-chain methyl groups provides the dominent source of relaxation in the first three; activation energies are 10.5 ± 1 1, 8.5 ± 1 kJ/mol, respectively, significantly lower than in the monomeric crystals. Relaxation times in poyglycine are two orders of magnitude longer than in the monomeric crystals. Relaxation times in polyglycine, significantly lower than in the monomeric crystals. Relaxation times in polyglycine are two orders of magnitude longer and are attributed mainly to segmental motions of the polymer chains. Evidence of nonexponential recovery of nuclear magnetization was encountered in the first three homopolyamino acids but not in polylycine, and was attributed to the correlated time to characterize these motions gave quite good agreement with the data; some improvement was obtained for two polymers using a Cole-Davidson distribution of correlation times. For biopolymers using a Cole-Davidson distribution of correlation times. For biopolymers generally it is concluded that rapid methyl group reorientation is a common dynamical feature and an important source of nuclear magnetic relaxation.     

Structural transitions in poly(A), poly(C), poly(U), and poly(G) and their possible biological roles

The homopolynucleotide (homo-oligonucleotide) tracts function as regulatory elements at various stages of mRNAs life cycle. Numerous cellular proteins specifically bind to these tracts. Among them are the different poly(A)-binding proteins, poly(C)-binding proteins, multifunctional fragile X mental retardation protein which binds specifically both to poly(G) and poly(U) and others. Molecular mechanisms of regulation of gene expression mediated by homopolynucleotide tracts in RNAs are not fully understood and the structural diversity of these tracts can contribute substantially to this regulation.

This review summarizes current knowledge on different forms of homoribopolynucleotides, in particular, neutral and acidic forms of poly(A) and poly(C), and also biological relevance of homoribopolynucleotide (homoribo-oligonucleotide) tracts is discussed. Under physiological conditions, the acidic forms of poly(A) and poly(C) can be induced by proton transfer from acidic amino acids of proteins to adenine and cytosine bases. Finally, we present potential mechanisms for the regulation of some biological processes through the formation of intramolecular poly(A) duplexes.     

Preresonance Raman studies of poly(L-lysine), poly(L-glutamic acid), and deuterated N-methylacetamides

The Raman spectra of poly(L -lysine) with various structures, ionized poly(L -glutamic acid), and deuterated N-methylacetamides have been observed using visible and the 257.3-nm laser lines as the light source. Most of the Raman bands with significantly enhanced intensities in the uv-excited spectra of the polymers have been assigned to the vibrations associated with the C?O and C–N stretching modes, the amide I, II, III, I′, II′, and III′, with reference to the results obtained for simple amide molecules including the deuterated N-methylacetamides. Several amide frequencies have been newly identified and the structures of the polymers have been discussed through the comparison of the Raman and ir amide frequencies.     

Energetics of Z-DNA formation in poly d(A-T), poly d(G-C), and poly d(A-C) poly d(G-T).

The conformational change for the alternating purine-pyrimidine polydeoxyribonucleotides i.e. poly d(A-T), poly d(G-C), and poly d(A-C) poly d(G-T) from a right-handed conformation at room temperature to the left-handed Z-DNA like double helix at elevated temperatures has been studied by UV spectroscopy, Raman spectroscopy, and by adiabatic differential scanning microcalorimetry (DSC) in the presence of Na+ and Mg2+ or Ni2+ respectively as counterions. The differential UV spectra reveal through a hyperchromic shift at around 280nm and a hypochromic shift at 260nm that a conformational change to the left-handed conformation occurs. The Raman spectra clearly show characteristic changes, a drastic decrease of the band at 680cm-1 and the appearance of a new band at 628cm-1, due to the change of the purine bases to the syn conformation upon inversion of the helix-handedness. The course of the transition as function of temperature can be followed quantitatively by plotting the change in the excess heat capacity vs. temperature. The transition enthalpy delta H for the B- to Z-DNA transition per mole base pairs (mbp) amounts to 2.0 +/- 0.2kcal for poly d(G-C), to 4.0 +/- 0.4kcal for poly d(A-T), and to 3.1 +/- 0.3kcal for poly d(A-C) poly d(G-T). The enthalpy change due to the Z-DNA to coil transitions (per mole base pairs) amounts to 11kcal for poly d(G-C), 10.5kcal for poly d(A-T) and 11.3kcal for poly d(A-C) poly d(G-T).     

Synthesis and coding properties of poly(c 1 A), poly(c 3 A), poly(c 7 A) and poly(h 6 A)

The homopolynucleotides poly(c¹A), poly(c³A), poly(c⁷A) and poly(h⁶A)⁺⁺ were synthesized from their corresponding nucleoside diphosphates using polynucleotide phosphorylase. With the exception of poly(h⁶A), which displayed no hypochromicity, the homopolynucleotides showed melting profiles similar to poly(A). All these polynucleotides, poly(h⁶A), poly(c⁷A), poly(c³A) and poly(c¹A) stimulated the binding of Lys-tRNA to ribosomes; the coding activity of poly(c¹A), however, was very low. Poly(h⁶A) was found to be less specific for Lys-tRNA than poly(A). The data supports the exclusive formation of Watson-Crick type base pairs and contradicts Hoogsteen base pairing in codon-anticodon recognition. Since, however, poly(h⁶A), which can form only one hydrogen bridge per base pair, stimulated the binding of Lys-tRNA comparably to poly(A), the coding activity of the homopolynucleotides tested is discussed in respect to their secondary structure as well as to the pK-values of their 6-amino groups.     

Recognition of polynucleotides by antibodies to poly(I), poly(C).

The binding of anti poly(I). poly (C) Fab fragments to double or triple stranded polynucletides has been studied by fluorescence. Association constants were deduced from competition experiments. The comparison of the association constants leads to the conclusion that several atoms of the base residues do not interact with the amino acid residues of the binding site of Fab fragment while the hydroxyl groups of furanose rings interact. These results suggest that the Fab fragments do not bind to the major groove of the double stranded polynucleotides. An interaction between the C(2)O group of pyrimidine residues and Fab fragments cannot be excluded. Circular dichroism of poly(I). poly(C) or poly(I). poly(br5C)-Fab fragments complexes are very different from the circular dichroism of free polynucleotides which suggests a deformation of the polynucleotides bound to the Fab fragments.     

CD studies of synthetic polypeptides as histone models: Poly(L-Arg-L-Ile-Gly), poly(L-Arg-L-Nva-Gly), and poly(L-Arg-L-Nle-Gly)

Sequential polypeptides (L -Arg-X-Gly)_n were prepared as synthetic models of arginine-rich histones to study their structure and their stereospecific interactions with DNA. In our previous work the conformational characteristics of poly(L -Arg-L -Ala-Gly), poly(L -Arg-L -Val-Gly), and poly(L -Arg-L -Leu-Gly) have already been analyzed. To obtain further insight into the influence of the X residue side chain on the conformation of the (L -Arg-X-Gly)_n polytripeptides, we now report their synthesis and cd properties when X represents the amino acid residues Ile, Nva, and Nle. The pentachlorophenyl active esters of the appropriate tripeptides were used to perform the polymerization, and the toluene-4-sulfonyl group was used to protect the arginine guanido group. CD spectroscopy showed that, in 100% trifluoroethanol, the degree of helical conformation increased in the order Ile → Nle → Nva. An equilibrium between β-turn, α-helix, and random-coil conformers occurred in 100% hexafluoroisopropyl alcohol, while a rise in the temperature or the addition of water favored the α-helix, the highest percentage of which was observed in a mixture of hexafluoroisopropyl alcohol: water (20 : 80) and in the order Ile → Nle → Nva. In aqueous solutions (at pH 7 and 12) the polymers behaved as a random coil, but they were forced to a less aperiodic structure, over a range of ionic strengths (0–0.5M NaF). A rise in temperature of up to 70°C in 100% trifluoroethanol resulted in a decrease of the α-helix percentage of the polymers, while in aqueous solutions the aperiodic structure decreased with increasing temperature. This study proved the importance of the nature of the X residue (length, C_β branching) in relation to the structural order of the sequential polypeptides. We concluded that the polymers prepared are suitable models for arginine-rich histones.     

High pressure fourier transform infrared spectroscopy of poly(dA)poly(dT), poly(dA) and poly(dT)

The effect of hydrostatic pressure upon the DNA duplex, poly(dA)poly(dT), and its component single strands, poly(dA) and poly(dT) has been studied by fourier-transform infrared spectroscopy (FT-IR). The spectral data indicate that at 28 degrees C and pressures up to 12 kbar (1200 MPa) all three polymers retain the B conformation. Pressure causes the band at 967 cm(-1), arising from water-deoxyribose interactions, to shift to higher frequencies, a result consistent with increased hydration at elevated pressures. A larger pressure-induced frequency shift in this band is observed in the single stranded polymers than in the double stranded molecule, suggesting that the effect of pressure on the hydration of single strands may be greater than upon a double stranded complex. A pressure-dependent hypochromicity in the bands attributed to base stacking indicates that pressure facilitates the base stacking in the three polymers, in agreement with previous assessments of the importance of stacking in the stabilization of DNA secondary structure at ambient and high pressures.     

Ferric complexes of acetoacetyl derivatives of poly(L-lysine), poly(L-ornithine), and poly(L-diaminobutyric acid). I. Preparation and absorption properties in solution

Poly(N^ε-acetoacetyl-L -lysine), poly(N^δ-acetoacetyl-L -ornithine) and poly(N^γ-acetoacetyl-L -diaminobutyric acid) form colored complexes with ferric ions in water/dioxane solutions. These complexes are soluble at pH values lower than 2.8 and show their maximum absorption at 257 nm in the uv and at 478 nm in the visible region; whereas the ferric complex of the model compound n-hexylacetoacetamide exhibits absorptions centered at 258 and 536 nm, respectively. It is shown that in the complex of the model compound one metal ion is bound per acetoacetamide group, while in the complexes of the three polymers two β-ketoamides side chains are bound per ferric ion under the same solvent, pH, concentration, and ionic strength conditions. The binding constants of ferric ions to the three polymers, and the formation constant of the ferric complex of the model compound are also evaluated.     

Metal complexs of poly (α-amino acids): Cu(II) chelates of acetoacetylated poly(L-lysine), poly(L-ornithine), and poly(L-diaminobutyric acid)

The cupric complexes of poly(N^ε-acetoacetyl-L -lysine), [Lys(Acac)]_n′ poly(N^δ-acetoacetyl-L -ornithine), [Orn(Acac)]_n′ and poly(N^γ-acetoacetyl-L -diaminobutyric acid), [A₂bu-(Acac)]_n, as well as of the model compound n-hexyl acetoacetamide, have been investigated by means of absorption, potentiometric, equilibrium dialysis, and CD measurements. While in the complex of the model compound, one chelating group is bound to one cupric ion, in the polymeric complexes two β-ketoamide groups are bound to Cu(II) under the same experimental conditions. The binding constant of cupric ions to the three polymers and the formation constant of the Cu(II)-nhexylacetoacetamide complex have been evluated. Investigation on the chiroptical properties of the three polymeric complexes shows that the peptide backbone does not undergo conformational transitions, remaining α-helical when up to 20% of the side chains are bound to Cu(II). The optical activity of the β-ketoamide chromophores is substantially affected by complex formation and is discussed in terms of asymmetric induction from the chiral backbone.     

Poly(gamma-L-diaminobutanoic acid), a novel poly(amino acid), coproduced with poly(epsilon-L-lysine) by two strains of Streptomyces celluloflavus

Two poly(ɛ- l -lysine) (ɛ-PL) producer strains of Streptomyces celluloflavus secreted a novel polymeric substance into their culture broths along with ɛ-PL. Three types of HPLC analysis plus one- and two-dimensional ¹H and ¹³C nuclear magnetic resonance experiments revealed that the secreted substance was poly(γ- l -diaminobutanoic acid) (γ-PAB), an l -α,γ-diaminobutanoic acid ( l -DAB) homopolymer linking between γ-amino and α-carboxylic acid functional groups. The γ-PABs from the two strains had an identical chemical structure, and the same number-average molecular weight of 2100–2200. No copolymers composed of the two amino acids l -DAB and l- lysine were found in either of the broths from the producers. Both strains coproduced high levels of the two poly(amino acid)s in the presence of SO₄²⁻ at pH 4.0 and 4.5 L min⁻¹ aeration in a 5-L jar fermentor. γ-PAB exhibited strong inhibitory activities against various yeasts and weaker actions against bacteria than ɛ-PL. γ-PAB may have various biological functions similar to ɛ-PL, and the use of γ-PAB along with ɛ-PL would be advantageous for technical applications in various fields.     

Heat capacities of solid poly(amino acids). I. Polyglycine,poly(L-alanine), and poly(L-valine)

Heat capacities of polyglycine, poly(L -alanine), and poly(L -valine) were analyzed using approximate group vibrations and fitting of the skeletal vibrations to a two-parameter (Θ₁, Θ₃) Tarasov function. New experimental data were measured by differential scanning calorimetry in the temperature range of 230–390 K. Good agreement between our experimental data and the calculated data was observed for all three poly(amino acids). Previous investigations showed agreement between calculation and reported experimental data for only limited low temperature ranges. At higher temperatures, discrepancies of up to 55% existed between experiment and calculation. The cause of this discrepancy must be assumed to be experimental error. Recommended experimental data are revised on the basis of this investigation. Computed heat capacities are available for the three biopolymers in the solid state from 0 to 1000 K.     

A study of conformational stability of poly(L-alanine), poly(L-valine), and poly(L-alanine)/poly(L-valine) blends in the solid state by (13)C cross-polarization/magic angle spinning NMR

13C cross-polarization/magic angle spinning (CP/MAS) NMR and (1)H T(1rho) experiments of poly(L-alanine) (PLA), poly(L-valine) (PLV), and PLA/PLV blends have been carried out in order to elucidate the conformational stability of the polypeptides in the solid state. These were prepared by adding a trifluoroacetic acid (TFA) solution of the polymer with a 2.0 wt/wt % of sulfuric acid (H(2)SO(4)) to alkaline water. From these experimental results, it is clarified that the conformations of PLA and PLV in their blends are strongly influenced by intermolecular hydrogen-bonding interactions that cause their miscibility at the molecular level.     

Syntheses and conformational studies of poly(Nε-methyl-L-lysine), poly(Nδ-methyl-L-ornithine), poly(Nδ-ethyl-L-ornithine), and their carbobenzoxy derivatives

The helix–coil transitions of poly(N^ε-methyl, N^ε-carbobenzoxy-L -lysine), poly(N^δ-methyl, N^δ-carbobenzoxy-L -ornithine), and poly(N^δ-ethyl, N^δ-carbobenzoxy-L -ornithine) in chloroform–dichloroacetic acid and their corresponding decarbobenzoxylated polypeptides in alkaline solutions were followed by optical rotation measurements. The introduction of a methyl or an ethyl group to the side chains of the carbobenzoxy derivatives of poly(L -lysine) and poly(L -ornithine) appeared to weaken the helical conformation at 25°C. The thermodynamic quantities of the three water-soluble polypeptides were calculated from the data on potentiometric titrations at several temperatures. For uncharged coil-to-helix transition, ΔH = ?370 cal/mol and ΔS = ?1.1 eu/mol for poly(N^ε-methyl-L -lysine), and ΔH = ?540 cal/mol and ΔS = ?1.6 eu/mol for poly(N^δ-ethyl-L -ornithine) (all on molar residue basis). The absolute values of ΔH and ΔS dropped in the region of pH-induced transition and eventually both quantities became positive. The initiation factor σ was about 2 × 10^?3, which was essentially independent of temperature. For poly(N^δ-methyl-L -ornithine) the coil-to-helix transition was not complete even when the polymer was uncharged at high pH.     

Vacuum UV CD of the low-salt Z-forms of poly(rG-dC).poly(rG-dC), and poly(dG-m5dC).poly(dG-m5dC)

The vacuum CD spectra of poly(rG-dC)·poly(rG-dC) and poly(dG-m⁵dC)·poly(dG-m⁵dC) have been obtained for the low-salt Z-conformations of both polymers. The spectra are very similar to those for the high-salt Z-forms. This behavior is consistent with the suggestion that the low- and high-salt Z-forms are comprised of different proportions of Z_I- and Z_II-conformations.     

Acoustical investigation of poly(dA).poly(dT), poly[d(A-T)], poly(A).poly(U) and DNA hydration in dilute aqueous solutions.

Apparent molar adiabatic compressibilities and apparent molar volumes of poly[d(A-T)].poly[d(A-T)], poly(dA).poly(dT), DNA and poly(A).poly(U) in aqueous solutions were determined at 1 degree C. The change of concentration increment of the ultrasonic velocity upon replacing counter ion Cs+ by the Mg2+ ion was also determined for these polymers. The following conclusions have been made: (1) the hydration of the double helix of poly(dA).poly(dT) is remarkably larger than that of other polynucleotides; (2) the hydration of the AT pair in the B-form DNA is larger than that of the GC pair; (3) the substitution of Cs+ for Mg2+ ions as counter ions results in a decrease of hydration of the system polynucleotide plus Mg2+, and (4) the magnitude of this dehydration depends on the nucleotide sequence; the following rule is true: the lesser is a polynucleotide hydration, the larger dehydration upon changing Cs+ for Mg2+ ions in the ionic atmosphere of polynucleotide.     

Ferric complexes of acetoacetyl derivatives of poly(L-lysine), poly(L-ornithine), and poly(L-diaminobutyric acid). II. Conformational properties in solution. Evidence for a stereospecific complex formation

The conformational properties of ferric complexes of poly(N^ε-acetoacetyl-L -lysine), poly(N^δ-acetoacetyl-L -ornithine), and poly(N^γ-acetoacetyl-L -diaminobutyric acid) were investigated in 1:1 water/dioxane by CD techniques. Optical activity was found in the visible and in the uv absorption region of the polymeric complexes. The conformation of the peptide backbone was always that of a right-handed α-helix, and was found independent of the degree of complexation, at least up to a degree of binding of 20%. In the absorption region of the side-chain chromophores the optical activity is substantially affected by complex formation. In all three cases a splitting of the ligand π → π* transition centered at 257 nm is observed. These data suggest a stereospecific complex formation. From the signs of the splitting it also appears that the chirality of the poly(N^δ-acetoacetyl-L -ornithine) complex is opposite that of the other two polymers.