bluuebunny/crossref_metadata_2025 huggingface dataset:
From synthetic antigens to synthetic vaccines
Authors: Sela Michael
Published: December 2004

The availability of synthetic antigens permitted a systematic elucidation of the molecular basis of antigenicity, as well as of other phenomena, and to establish the genetic control of immune response and its link to the major histocompatibility region of the species. Moreover, it allowed a conceptual approach to production of vaccines made up of synthetic fragments of relevant virus antigens. This was first demonstrated with a bacteriophage, MS‐2, that can be efficiently inactivated with antibodies obtained with an immunogen in which a synthetic peptide corresponding to a sequence of 20 amino acid residues within its coat protein was covalently linked to a polymeric synthetic carrier. When the synthetic adjuvant N‐acetyl‐muramyl‐L‐alanyl‐D‐isoglutamine (MDP) was chemically linked to the same molecule, the resulting immunogen was effective in aqueous solution. This approach has now been used for diphtheria: a synthetic peptide from the A‐chain of the toxin, covalently attached together with MDP to the synthetic carrier multichain poly(DL‐alanine) induced antibodies cross‐reactive with the diphtheria toxin as well as protection against the dermonecrotic activity of the toxin. A similar approach has been described also by R. Arnon and her colleagues in our laboratory for influenza virus, making use of a peptide with sequence 91–108 of the hemagglutinin of type A H3N2 strains attached to tetanus toxoid. The results indicated that the conjugate led to partial protection against A/Texas mouse‐adapted influenza virus.
Contribution of synthetic polymers of amino acids to knowledge of immune response
Authors: Maurer Paul H. , Babu Uma Mahesh , Lai Chang‐Hai
Published: December 2004

The synthetic random polymers poly(Glu,Lys,Phe), poly(Glu,Phe) and poly(Glu,Lys,Tyr), have been used to study some parameters associated with the genetic control of the immune response (Ir) of mice. Mice of haplotypes d and q respond well to GLPhe. Mice of haplotypes k and b were previously shown to be nonresponders, whereas the F1 (k × b) responded via a phenomenon involving “complementation” between 2 Ir genes, i.e., one gene product from IA, and another from IE form the requisite two‐chain Ia “receptor” macromolecules (EE). When it was determined that mice of haplotypes q and k respond to GPhe, and the controlling gene maps to IA, (Aα Aβ), we tested the theory that mice having q and k alleles in IA might respond to GLPhe via recognition of GPhe determinants in the terpolymer. Employing the in vitro proliferative response to T‐cells from mice immunized with GLPhe and stimulated with GPhe and GLT (cross‐reaction), it was determined that different determinant selection patterns exist in the recognition of GLPhe. Mice having q and k alleles in IA can respond to GLPhe via one mechanism, and other mice having d and f alleles respond via other mechanisms. The F1 of the appropriate nonresponder strains forming the Ia molecule (EE) still exhibit the “complementation” phenomenon. Rabbit antibody against anti‐GPhe (ID) from SWR mice (H‐2q)(anti‐ID) was prepared. This anti‐ID strongly inhibited the binding of 125I‐GPhe by anti‐GPhe antisera produced only in mice of H‐2q haplotype and had no effect on the binding of GPhe by anti‐GPhe antisera produced in mice of other haplotypes. The anti‐ID also inhibited the binding of 125I‐GLPhe and 125I‐GPhe by anti‐GLPhe antisera produced only in mice of H‐2q haplotype. These specificities were also confirmed by the inhibition of the plaque‐forming cells. It was concluded that the antibodies produced in mice of H‐2q haplotype against GPhe and GLPhe share common idiotypic determinants that are recognized by the anti‐idiotypic antiserum.
The T‐cell receptor as analyzed by functional T‐cell lines specific to a synthetic polypeptide antigen
Authors: Mozes Edna
Published: December 2004

For a better understanding of the molecular nature of the antigen‐specific T‐cell recognition system, continuous T‐cell lines specific to the synthetic polypeptide antigen poly(Tyr,Glu)‐poly(DLAla)‐‐poly(Lys) [(T,G)–A‐‐L] were established from C3H.SW (high‐responder) activated T‐cells, cloned, and characterized. These lines and their derived clones are also constitutive secretors of antigen‐specific T‐cell replacing helper factors. The secreted T‐cell helper factor was shown to possess MHC determinants as well as V‐region determinants, or more specifically, idiotypic determinants that are cross‐reactive with those expressed on (T,G)–A‐‐L‐specific antibodies of the same mouse strain. Using the fluorescence‐activated cell sorter (FACS II) and individual C57BL/6 anti‐idiotypic sera produced against (T,G)–A‐‐L‐specific antibodies of C3H.SW origin, we have demonstrated the expression of the cross‐reactive idiotypic markers on the monoclonal helper T‐cells. Attempts were made to purify the active fraction of the T‐cell factors secreted by the (T,G)–A‐‐L continuous helper lines. Gel analysis of the twice affinity‐purified eluate of a (T,G)–A‐‐L column revealed the existence of iodinated bands with molecular weight of 17,000 and 15,000, in addition to a diffuse band of high molecular weight. The specific helper activity of the factors was associated with a 65–75% ammonium sulfate precipitate. Gel electrophoresis of the latter fraction, as well as of an eluate of a (T,G)–A‐‐L–Sepharose column indicated that a high‐molecular‐weight (< 67,000) and a low‐molecular‐weight (15,000–17,000) fraction contained the biological activity of the factor. Similar results were obtained following chromatography of the factor on Sephadex G‐100 columns. The two fractions were shown to be synthesized by the T‐cell lines, as indicated by internal labeling experiments using 35S‐methionine. Thus, it is suggested that a fraction of an apparent molecular weight of 15,000–17,000 preserves both the antigen specificity and the helper activity of the factor produced by the (T,G)–A‐‐L‐specific T‐cell lines.
Some novel approaches to the design and synthesis of peptide–catecholamine conjugates
Authors: Verlander M. S. , Jacobson K. A. , Rosenkranz R. P. , Melmon K. L. , Goodman M.
Published: December 2004

A series of novel, functionalized catecholamines (congeners) has been synthesized in which, formalistically, the N‐isopropyl group of isoproterenol has been extended by a linear alkyl chain of varying length, terminated by a carboxyl group. Model amide derivatives have also been prepared in order to optimize the biological activity of these derivatives and also to aid in the design of appropriate peptides for the synthesis of conjugates. As a result of these studies, a series of amino acid and monodisperse peptide carriers, containing p‐aminophenylalanine as the point of attachment for the drug, was prepared, together with the corresponding conjugates. In vitro and in vivo evaluation of the congeners, model amides, and conjugates has demonstrated that the biological activity of these derivatives is extremely sensitive to structural modifications at a point far‐removed from the pharmacophore, in both the congener amide and conjugate series. A number of the model amides and conjugates have proven to be highly active when tested in both in vitro and in vivo test systems. The implications of these results in terms of a novel structure–activity approach to drug design are discussed.
Synthesis and biological activity of (D)Ala2, Leu5‐enkephalins containing hydrophilic or hydrophobic moieties
Authors: Filippi Bruno , Biondi Laura , Filira Fernando , Rocchi Raniero , Bellini Cecilia , Sarto Giampietro
Published: December 2004

The synthesis in solution of some modified (D)Ala2,Leu5‐enkephalins has been carried out. The lipophilic properties of the parent compound have been modified by amidation of the carboxyl function with alkylamines of increasing hydrophilicity to increase permeability of the blood‐brain barrier. Attempts to reduce enzymatic degradation have been carried out either by reductive glucosamination or by amidation of the carboxyl function with 2‐amino‐2‐deoxy‐β‐D‐glucopyranose. Esterification of the carboxyl function of (D)Ala2,Leu5‐enkephalin with polyethylenglycole 1000 has also been carried out. The effects induced by these modifications have been evaluated by in vitro and in vivo tests.
Centrifugal field relaxation and ionic strength effects on calf thymus DNA gels
Authors: Richard Alfred J.
Published: December 2004

The gellike phase formed by DNA on sedimentation to the bottom of a centrifuge cell has been studied as a function of centrifuge speed and of solvent ionic strength. The swelling pressure of the gel was found to be logarithmically related to the DNA concentration and also to the ionic strength of the solvent. The concept of electrostatic persistence length was useful in interpreting the gel‐phase volume changes occurring with ionic strength.
1H‐nmr studies on the dinucleoside monophosphates containing 2′‐halogeno‐2′‐deoxypurinenucleosides: Effects of 2′‐substitutes on conformation
Authors: Uesugi Seiichi , Kaneyasu Toshinori , Imura Junko , Ikehara Morio , Cheng Doris M. , Kan Lousing , Ts'o Paul O. P.
Published: December 2004

Seven dinucleoside monophosphates containing 2′‐halogeno‐2′‐deoxypurine nucleoside residue, dAfl‐U, dAcl‐U, dAbr‐U, dAio‐U, dGfl‐U, and dIfl‐C, were chemically synthesized and investigated by 1H‐nmr spectroscopy at 300 MHz. The sugar and backbone conformations of these compounds were analyzed by the spectral pattern of furanose proton resonances; and the extents of base‐base interaction were estimated from chemical shifts and their temperature‐dependent changes of base‐proton resonances. It is found that the population of C3′‐endo conformer and the extent of base‐base interaction decrease as the electronegativity of 2′‐substituent decreases in dAx‐U (x = fl, cl, br, and io) series. The C3′‐endo (3E) population and the base‐base interaction in Nfl‐U (N = A,G)‐type dimers as well as dIfl‐C are relatively higher than the corresponding natural ribo‐dimers but can be recognized as grossly similar to the conformation of regular RNA dimers.
Conformation of sequential and random copolypeptides of lysine and alanine in sodium dodecyl sulfate solution
Authors: Kubota Shigeo , Ikeda Kiyoshi , Yang Jen Tsi
Published: December 2004

A series of sequential polypeptides, (Lysi‐Alaj)n, and random copolypeptides, (Lysx, Alay)n, were synthesized. The competitive effect of Ala, a helix former, and Lys, whose homopolymer has a β‐form in neutral NaDodSO4 solution, was determined by CD and absorption spectroscopy. All the polypeptides studied were unordered in neutral solution without the surfactant. Of the six sequential polypeptides only (Lys‐Ala)n adopted a stable β‐form in NaDodSO4 solution. Most striking is the difference between this polypeptide, (Lys2‐Ala2)n and (Lysx, Alay)n, even though they all have equimolar Lys and Ala. (Lys2‐Ala2)n was partially helical in 2.5–5 mM NaDodSO4 but approached the unordered form in 50 mM NaDodSO4, whereas (Lys50, Ala50)n was completely helical in all NaDodSO4 concentrations. Even Lysrich (Lys2‐Ala)n and (Lys3‐Ala)n formed a partial helix and a trace of the β‐form, respectively, in low NaDodSO4 concentrations; both reverted to the unordered form in high NaDodSO4 concentrations. These results can be explained by Pauling‐Corey's model for β‐pleated sheets. Only (Lys‐Ala)n has all DodSO‐bound Lys+ residues on one side and Ala residues on the other side of the polypeptide chain. They can nestle quiet efficiently in a β‐sheet and between neighboring β‐sheets. Our results further imply that random copolypeptides are not completely random; they comprise varying segments of (Lysk‐Alam), where k and m could vary from zero to a small integer.
Complex formation with alkali and alkaline earth metal ions of cyclic octapeptides, cyclo(Phe‐Pro)4, cyclo(Leu‐Pro)4, and cyclo[Lys(Z)‐Pro]4
Authors: Kimura Shunsaku , Imanishi Yukio
Published: December 2004

Complex formation with alkali and alkaline earth metal ions of cyclic octapeptides, cyclo(Phe‐Pro)4, cyclo(Leu‐Pro)4, and cyclo[Lys(Z)‐Pro]4 was investigated in relation to conformation. In an alcohol solution, cyclo(Phe‐Pro)4 did not form complexes. However, cyclo(Leu‐Pro)4 and cyclo[Lys(Z)‐Pro]4 formed complexes selectively with Ba2+ and Ca2+ ions. Changing the solvent from alcohol to acetonitrile, the complexation behavior was very different. In acetonitrile, cyclo(Phe‐Pro)4 was found to form a complex with Ba2+, and CD spectra of cyclo(Leu‐Pro)4 and cyclo[Lys(Z)‐Pro]4 changed sharply on complexation with K+. Rate constants of the complex formation between the cyclic octapeptides and metal salts were in the range of 0.7–12 L mol−1 min−1 in an alcohol solution. One of the two types of complex formation in acetonitrile was much faster than that in an alcohol solution.
Heat capacity increments: Conformational transitions in polypeptides
Authors: Couchman P. R. , Gajnos G. E. , Ryan C. L. , Karasz F. E.
Published: December 2004

A thermodynamic treatment of the helix–coil transition of synthetic polypeptides in binary organic solvent mixtures is extended to describe isobaric heat‐capacity increments associated with the phenomenon. This development resolves such increments into three components: two associated respectively with intrinsic differences between the ordered and disordered states of the macromolecule and between the coil–solvent complex and its components, and a third term derived from the temperature dependence in the fraction of coil residues bound to active solvent. Insights derived from this analysis are also applied to the discussion of some heat capacity increments associated with the denaturation of globular proteins.
Low‐frequency Raman scattering by anharmonic DNA modes with A⇌B character modeled with a classical double‐well oscillator
Authors: Martin A. J.
Published: December 2004

The optical modes of DNA that displace one strand against the other are modeled by the motion of an oscillator in an asymmetric quartic double‐well potential whose minima represent the A‐ and B‐conformations. Assuming that the variation of the polarizability during vibration derives mainly from the tilting of the base rings relative to the helix axis, the total polarizability tensor is shown to possess approximately ellipsoidal symmetry and to depend nonlinearly on the instantaneous displacement of the two strands. The Raman spectrum of a collection of randomly oriented molecules is calculated. It consists of one or more peaks with characteristic shape. The depolarization ratio is 3/4, independent of molecular conformation and frequency. The results are discussed in the light of existing experimental and theoretical information.
31P‐NMR studies of deoxyoligonucleotides
Authors: Cheng Doris M. , Kan Lou‐Sing , Iuorno Virginia L. , Ts'O Paul O. P.
Published: December 2004

The assignments of 31P resonances of eight short oligonucleotides have been achieved through specific heterodecoupling techniques reported previously from our laboratory [Cheng et al. (1982) Biopolymers 21, 697–701]. The temperature dependence of the assigned 31P chemical shifts of these oligomers was studied and compared to the constitutive dimeric units. This comparison gives some insight to the chemical‐shift values of the phosphorus resonances of oligonucleotides. (1) The 3′‐end terminal phosphorus resonance in an oligomer tends to locate at a spectral position relatively close to its constitutive dimeric unit. (2) On chain elongation (from 5′‐toward 3′‐end), the phosphorus resonance in the oligomer will be shifted upfield by 0.2–0.3 ppm, as compared to its constitutive dimeric unit. (3) The relative positions of phosphorus resonances in an oligomer tend to remain in the same order as their constitutive dimeric units. The above obsrvations lead to the development of a new procedure to assign the 31P resonance of a single‐stranded oligonucleotide.
Conformational studies of adenosine 5′‐phosphorodiamidates in aqueous solution
Authors: Tomasz Jenö , Barabás Klára , Bottka Sándor , Kajtár Judit , Kajtár Márton , Kajtár‐Peredy Mária , Radics Lajos
Published: December 2004

CD and nmr studies show that the conformational equilibrium of N(P)‐alkylated adenosine 5′‐phosphorodiamidates 2–4 and, to a much lesser extent, adenosine 5′‐phosphorodiamidate 1 differ at two parts from that of adenosine 5′‐phosphate in water. The glycosidic bond conformation is slightly shifted into the direction of the syn range, and the ribose ring N‐conformer population increases. The conformational changes increase with increasing degree of N(P)‐alkylation and cause a change of the sign of the long‐wavelength CD band for 2–4. The new conformation may be characterized by a stacked arrangement for the adenine ring and the 5′‐substituted phosphate group of 2–4. Hydrophobic attraction between the head and the tail of the molecules, as well as the entropy effect (which promotes the clustering of apolar faces in water), may be jointly responsible for this stacked arrangement. The intramolecular attractive forces stabilizing the stacked conformation may act at the expense of the solubility decreasing intermolecular attractive forces, and may thus be responsible for the increased water solubility of 2–4, which is higher than that of 1 by more than two orders of magnitude.
Aggregation of poly(γ‐benzyl‐α,L‐glutamate)
Authors: Chakrabarti Sumana , Miller Wilmer G.
Published: December 2004

The aggregation of poly(γ‐benzyl‐α,L‐glutamate) and its enantiomer in toluene has been investigated by following the viscosity as a function of temperature, concentration, molecular weight, molecular‐weight distribution, helix chirality, and shear rate. The temperature and concentration data for a 138,000‐molecular‐weight sample was fitted to an open, reversible end‐to‐end aggregation model. The aggregation numbers resulting from this fit were consistent with the sudden onset in non‐Newtonian flow resulting from only a 0.2‐wt% increase in concentration. The association equilibrium constant was then used to predict viscosity for comparison with other data, in particular, the effect of molecular weight and molecular‐weight distribution. A mixture of right‐and left‐handed helices showed the aggregation was not chiral selective. The stiffness of end‐to‐end aggregated (hydrogen‐bonded) molecules differed little from their covalent counterparts, at least below a molecular weight of ∼106. We conclude that polybenzylglutamate aggregation in toluene can be described by an open end‐to‐end aggregation model.
The titratable joint phenomenon in ΦW‐14 DNA
Authors: Wilcoxon Jess , Schurr J. Michael , Warren R. A. J.
Published: December 2004

Dynamic light scattering (DLS) studies are carried out on native ΦW‐14 DNA, which has putrescines covalently attached at the methyl groups of half its thymines, and on a chemically modified form of the same DNA in which the ammonium groups of its putrescines are almost completely acetylated. From neutrality to pH 9.6, both forms of ΦW‐14 DNA exhibit the same curve of Dapp vs K2 over the range K2 = 0.5 × 1010 to K2 = 20 × 1010 cm−2, and this coincides with curves that we have observed for other DNAs. (Dapp, apparent diffusion coefficient; K, scattering vector). However, when the pH is raised to pH 10.0–10.2, native ΦW‐14 exhibits a spectacular decrease in Dapp at large wave vector, whereas the acetylated form shows no sign of such behavior. It is inferred that bound ammonium groups make an essential contribution to the stabilization of titratable joints. Comparing the pH profiles of the absorbance (A260) for these two DNAs gives some evidence that base unstacking may be involved in titratable joint formation.
Interaction of poly‐5‐bromouridylic acid. II. Thermodynamic analysis of its interaction with adenosine and 9‐methyladenine
Authors: Higuchi Shigesada , Yasui Kumiko
Published: December 2004

The interaction of poly‐5‐bromouridylic acid [poly(BU)] with adenosine and 9‐methyladenine was studied by equilibrium dialysis, optical melting, and microcalorimetry. The stacking free energy, ω, was estimated as −17.6 kJ/mol for adenosine·2poly(BU) and −18.8 kJ/mol for 9‐methyladenine·2poly(BU) from the binding isotherms constructed from equilibrium dialysis results. The binding isotherms constructed from a series of melting curves also gave ω values for adenosine·2poly(BU). The thermal stability of the complex depends on monomer concentration, and the partial molar enthalpies of the complex formation at the midpoint of the transition were evaluated from the Tm coefficients as a function of free monomer concentration. The values of −92.0 and −90.4 kJ/mol were obtained for adenosine·2poly(BU) and 9‐methyladenine·2poly(BU) in 0.4M NaCl–0.02M Na‐cacodylate–5 × 10−4M EDTA (pH 7.0), respectively. Microcalorimetric measurements provided lower integral heats of reaction values for these complexes, i.e., −73.2 kJ/mol for adenosine·2poly(BU) and −71.5 kJ/mol for 9‐methyladenine·2poly(BU). A comparison with a polyribouridylic acid system provided a quantitative understanding of a stabilization by bromination in terms of thermodynamic parameters.
Proton‐NMR study of the interaction of trans‐dichloro‐diamine‐platinum(II) with poly(I) and poly(I)·poly(C)
Authors: Fazakerley G. Victor , Hermann Dominique , Guschlbauer Wilhelm , Hawkes Geoffrey E.
Published: December 2004

The fixation of trans‐(NH3)2Cl2 Pt(II) to poly(I)·poly(C) at low rb (< 0.05) leads to the formation of two complexed species. The major species (ca. 82% of bound platinum) involves coordination of platinum to a single hypoxanthine base, while the other species involves coordination of two hypoxanthine bases, which are either far apart on the same strand or on separate poly(I) strands, to the platinum. These same two species are found after reaction with poly(I), as are two other species throughout the entire rb range studied (rb = 0–0.30). The latter two species are assigned to trans‐Pt bound to two bases on a poly(I) strand with (a) one or (b) two free bases between the two bound bases. These two species, (a) and (b), account for ca. 35% of the bound platinum, although the 1:1 species remains dominant (ca. 55%). These two additional species are observed at high rb (>0.075) after reaction with poly(I)·poly(C) but as very minor species. They are formed by reaction with melted poly(I) loops. Also at high rb, we have observed a shifted cytidine H5 resonance arising from interaction of trans‐Pt with a melted loop of poly(C). Most probably, this arises from an intramolecular poly(I) to poly(C) crosslink. Results from the reaction of trans‐Pt with poly(C) are presented for comparison.
Monoazido analog of ethidium as a chromatin probe: Binding to DNA
Authors: Coffman George L. , Yielding Lerena W. , Yielding K. Lemone
Published: December 2004

Two photoaffinity analogs of ethidium, 8‐azido‐3‐amino, and 3‐azido‐8‐amino‐5‐ethyl‐6‐phenylphenanthridinium chloride, have been used to probe the structure of mammalian chromatin and its interactions with the ethidium moiety. The monoazido analogs were established as suitable probes by comparing their interactions with chromatin and pure DNA prepared from chromatin to those of the parent ethidium bromide. Scatchard analysis of the binding data determined from spectrophotometric titrations showed that the analogs interacted with both nucleic acids in a manner similar to the parent compound. The effect of chromatin proteins on the interaction of the ethidium moiety with intact chromatin was investigated directly. By exposing the noncovalent complex to visible light, the monoazido analog was attached covalently in its interaction sites within chromatin, and the amount of drug bound covalently to DNA was determined for both protein‐free DNA and chromatin. Using saturating concentrations of drug, DNA within intact chromatin was found to be associated with only half as much drug as DNA extracted from its protein prior to drug exposure. The distribution of drug bound within chromatin was determined following the attachment of the monoazido analog (by photoactivation) to chromatin that had undergone limited nuclease digestion. Several distinct populations isolated by size fractionation and quantitative measurements revealed that (1) both the core particles and the spacer‐containing particles contained bound drug, reflecting high‐affinity binding sites; and (2) chromatin particles containing 150 DNA base pairs (putatively nucleosome core structures) contained less total bound drug at high drug concentrations than those particles having intact spacer DNA.
Premelting and the hydrogen‐exchange open state in synthetic RNA duplexes
Authors: Preisler R. S. , Mandal C. , Englander S. W. , Kallenbach N. R. , Frazier J. , Miles H. T. , Howard F. B.
Published: December 2004

Here we attempt to relate equilibrium temperature‐dependent spectral changes in two synthetic RNA homopolymer duplexes—poly(rA) · poly(rU) and poly(rI) · poly(rC)—to the conformational opening detected in stopped‐flow hydrogen–deuterium exchange experiments on these molecules. We are concerned with changes in several spectral properties that occur well below onset of the thermally induced helix‐coil “melting” transition in these systems. These are known as “premelting” transitions, and can be detected in uv CD spectra as well as in vibrational bands of the bases in the ir. Both CD and ir spectra exhibit isoelliptic or isosbestic points consistent with a well‐defined two‐state premelting process. Application of a least‐squares analysis to two‐state models for premelting using data from different bands in the CD and ir shows that the enthalpies are substantially greater than that of the hydrogen‐exchange opening. Thus the hydrogen‐exchange open state represents only one premelting reaction among several that lead to equilibrium changes in helix geometry or base vibrational modes. The latter include processes that occur on a rapid time scale, including potential base‐pair openings not productive for the exchange reaction. It appears that the former, and not the hydrogen‐exchange opening, dominates the premelting alterations monitored by ir and CD spectroscopy.
Proton magnetic resonance studies on dermorphin and its peptide fragments
Authors: Pastore A. , Temussi P. A. , Tancredi T. , Salvadori S. , Tomatis R.
Published: December 2004

Dermorphin (TyrD‐AlaPheGlyTyrProSerNH2), a potent natural peptide opioid, its synthetic L‐Ala2 analog, and all the N fragments from the tripeptide (TyrD‐AlaPheNH2) to the parent hexapeptide amide were characterized for the first time by means of proton nmr spectroscopy at 11.74 T. Assignments of most protons of dermorphin were facilitated by the study of the N‐terminal fragments. Comparison of spectroscopic parameters with relative pharmacological activity is proposed as a possible means of studying flexible agonists in solution.
Generalized Poisson‐Boltzmann calculation of the distribution of electrolyte ions around the B‐ and Z‐conformers of DNA
Authors: Pack George R. , Klein Brian J.
Published: December 2004

A solution to the three‐dimensional Poisson‐Boltzmann equation, generalized to include the finite size of the ions, is presented for the environment of DNA in the B‐ and Z‐conformations. The results clearly indicate that despite the lower linear charge density of the left‐handed Z‐conformer, there is a higher concentration of Na+ at the immediate surface of the Z‐from than at the surface of the B‐form. The average concentration of counterions within a 12‐Å radius of the DNA is, nonetheless, higher for the B‐ than for the Z‐form. Calculations of the electrostatic interactions of these conformers with an environment of 0.01M monovalent salt show that the salt exerts a greater stabilizing effect on the left‐handed conformer than on the right‐handed form.
Low‐frequency dynamics of solid poly(L‐alanine) from Raman spectroscopy
Authors: Tipping M. , Viras K. , King T. A.
Published: December 2004

Raman modes from amorphous α‐helical poly(L‐alanine) in the low‐frequency region < 150 cm−1 have been observed and assignments and values compared with mode‐analysis calculations. The temperature dependence of the complete Raman spectrum of α‐poly(L‐Ala) is also reported.
Synthesis and characterization of six sequential polypeptides containing tyrosine, glutamic acid, alanine, and glycine by circular dichroism and difference spectroscopy
Authors: Zeiger Allen R. , Ellis John S. , Maurer Paul H.
Published: December 2004

Six sequential polytetrapeptides containing equimolar amounts of tyrosine, glutamic acid, alanine, and glycine were characterized by CD and difference spectroscopy over a wide range of pH. As the pH was lowered from physiological values, each of the polymers underwent pH‐sensitive transitions. The CD spectra indicated that two polymers, poly(Tyr‐Glu‐Ala‐Gly) and poly(Tyr‐Ala‐Glu‐Gly), had some α‐helical conformation at pH 7.0 and approached maximum helicity around pH 6.0; two others, poly(Ala‐Tyr‐Glu‐Gly) and poly(Glu‐Ala‐Tyr‐Gly), had no α‐helical conformation at pH 7.0 and about one‐third of the ellipticities of the above two polymers at pH 5.5; and the remaining two, poly(Ala‐Glu‐Tyr‐Gly) and poly(Glu‐Tyr‐Ala‐Gly) had little or no α‐helix, even at pH 5.5. Difference spectroscopy at 286 nm yielded results quite different. The molar extinction coefficients for poly(Tyr‐Glu‐Ala‐Gly) and poly(Tyr‐Ala‐Glu‐Gly) continued to change, even below pH 5.5, and the total changes in absorbance between pH 8.0 and 4.5 were of intermediate magnitudes among the six polymers. Poly(Ala‐Tyr‐Glu‐Gly) and poly(Glu‐Ala‐Tyr‐Gly), which had similar CD spectra, had the lowest and highest pH‐related changes in the molar extinction coefficients. It thus appears that amino acid composition alone cannot account for the apparent differences in conformation among the polytetrapeptides. Other factors, such as amno acid sequence, must play a major role in the determination of conformation.

The intrinsic viscosity of poly(Tyr‐Glu‐Ala‐Gly) increased markedly between pH 6.0 and 5.5, which was below the pH of the CD transition but above the pH at which the largest absorption perturbation change, at 286 nm, took place. The model that can best account for the relatively low pH at which the absorption transition of tyrosine occurred is a progressive immobilization of side chains in the α‐helix as the pH decreases.
Stabilization of short helices by intramolecular cluster formation
Authors: Mattice Wayne L.
Published: December 2004

The intramolecular formation of multiple clusters of interacting helices has been characterized in a homopolymer. The configuration partition function permits the formation of clusters in which the number of interacting helices may be as large as the greatest integer in n/2, where n denotes the number of amino acid residues in the chain. The theoretical formulation has its origin in a recent [Mattice, W. L. & Scheraga, H. A. (1984) Biopolymers 23, 1701–1724], tractable matrix expression for the configuration partition function for intramolecular antiparallel β‐sheet formation. Reassignment of the expression for one of the n(n+3)/2 elements in the sparse statistical weight matrix, along with a simple change in notation, converts that treatment into a matrix formulation of the configuration partition function for a chain containing multiple clusters of interacting antiparallel helices. The five statistical weights used are δ, fl, w, and the Zimm‐Bragg σ and s. Each tight bend that connects two interacting helices contributes a factor of δ, fl is used in the weight for larger loops between interacting helices, and w arises from helix–helix interaction. The influence of the helix–helix interaction is well illustrated by two helix–coil transitions in a chain with n = 156 and σ = 0.001. In the absence of helix–helix interaction, the transition occurs by the nucleation and subsequent elongation of a small number of helices. When helix–helix interaction is attractive, the transition can occur by a different mechanism. Formation of a single pair of interacting helices is followed by addition of new helices to the initial cluster. In the latter process, individual helices experience relatively little growth after they are formed.
A molecular mechanical study of netropsin‐DNA interactions
Authors: Caldwell James , Kollman Peter
Published: December 2004

We present molecular mechanical calculations on the complexes of netropsin with dA6·dT6, d(TATATA)2, d(CGCGCG)2, and d(CGCGAATTCGCG)2. The complexes were model built using computer graphics and then completely energy refined. Our calculations are consistent with the observed AT preference for netropsin and suggest that mixed sugar pucker geometries should be more stable than uniform in netropsin complexes with poly[d(A‐T)]·poly[d(A‐T)] and poly(dA)·poly(dt). The netropsin·d(TATATA) and netropsin·dA6·dT6 complexes are significantly different in structure, leading to a possible reason why the observed thermodynamics of netropsin‐association with poly[d(A‐T)]·poly[d(A‐T)] and with poly(dA)·poly(dT) are so different. We also model built and energy refined a structure of netropsin‐d(CGCGAATTCGCG)2 using as a guide the nmr data of Patel [(1982) Proc. Natl. Acad. Sci. USA, 79, 6424–6428] and found a three‐dimensional structure qualitatively consistent with the NOE enhancements observed by him. After our calculations were completed, we learned of an x‐ray structure of a netropsin:d(CGCGAATTCGCG)2 complex, and we compared the structure found in our calculation with the x‐ray structure.
Self‐association and solubility of peptides: Solvent‐titration study of Nα‐protected C‐terminal sequences of substance P
Authors: Toniolo Claudio , Bonora Gian Maria , Stavropoulos George , Cordopatis Paul , Theodoropoulos Dimitrios
Published: December 2004

Self‐association of Nα‐protected peptides related to C‐terminal sequences of substance P in methylene chloride was disrupted by adding increasing amounts of various polar organic solvents. This process was monitored by the disappearance of the amide I CO stretching band (1630 cm−1) of strongly intermolecularly H‐bonded molecules in the irabsorption spectra. The effects induced by main‐chain length, incorporation at position 9 of a residue promoting folding (α‐aminoisobutyric acid), the nature of solvent, and peptide concentration were established. A corollary 1H‐nmr investigation provided detailed information on the NH protons involved in the self‐association process as H‐bonding donors. The increasing propensity to aggregate exhibited by these peptides is paralleled by a decrease in their solubility. The impact of these results on the synthesis of substance P short sequences is briefly outlined.
Dependence of oxygen quenching of intercalated methylene blue triplet lifetime on DNA base‐pair composition
Authors: Berkoff Bruce , Hogan Mike , Legrange Jane , Austin Robert
Published: December 2004

We utilize the dynamic quenching of the triplet state of methylene blue by molecular oxygen to observe changes in the rate at which oxygen can penetrate the helix as a function of base‐pair composition. The results indicate that the interior of the oligonucleotide dA‐dT is more accessible than dG‐dC to small molecules such as dioxygen.
The relation of ion pairs to protein hydration: An IR spectroscopic and X‐ray crystallographic survey
Authors: Poole P. L. , Barlow D. J.
Published: December 2004

We report here a novel ir technique that we use to determine the apparent van't Hoff's energy for the distillation of water from protein amide and acid groups. The method is used to study the hydration properties of ten different proteins. The results show that water removal from the environment of ion‐paired acid groups (those ⩽ 4 Å from a basic group on the protein surface) requires less energy than water removal from the environment of nonpaired acid groups. These observations are supported by an analysis of the solvent distributions in protein crystal structures. A model is proposed for the interaction between water and protein acid groups, and this model is used to account for the distributions and properties of certain eye‐lens proteins.
Conformational changes of pinellin in solution using intrinsic fluorescence and CD as probes
Authors: Lu Zi‐Xian , Shi Qing‐Luo , Xu Qing‐Yu
Published: December 2004

Pinellin is a plant protein extracted from the rhizome of the Chinese herb Pinellia. It has the ability to abort early pregnancy in mice as well as in rabbits. Studies on the conformational changes of pinellin have been carried out in our laboratory using intrinsic fluorescence and CD. Experimental results show that some tryptophanyl side chains are buried more deeply than others, which results in the heterogeneity of tryptophanyl emission. CD data indicated a high content of β‐pleated sheet and β‐turn for the backbone conformation. The results of fluorescence and CD measurements both demonstrated the presence of intermediates along the path of denaturation. The following was proposed as the unfolding mechanism of pinellin in 6M guanidine hydrochloride: native state → first intermediate → second intermediate → fully unfolded state.
A polynucleotide CD probe: Interactions with oligomers of a polycationic amine that exhibit positive extrinsic bands at > 300 nm
Authors: Zarlenga D. S. , Halsall H. B. , Day R. A. , Roth A. C.
Published: December 2004

A set of large positive extrinsic CD bands ([θ]333 = 2.6 X 104 deg‐cm2/decimole phosphate) in the > 300 nm region as well as diminution of the intrinsic signals (θ275) have been observed in the CD spectra of various nucleic acids complexed with the achiral compound, N‐poly{α‐[N‐(4‐pyridylethylene‐4‐pyridyl‐N′‐)α′‐p‐xylyl]dibromide}‐4‐pyridylethylene‐4‐pyridinium bromide, (polymer X).1,2,5 The signal changes are attributed to the binding of polymer X chromophores isogeometrically to the DNA helix in an ordered chiral arrangement. Fractionation of polymer X gives 10 well‐separated oligomers. The oligomers were characterized by nmr. Their interactions with DNA have been investigated with respect to r(r = ratio of equivalents of polymer X charge/g‐atoms DNA phosphorus) and n (oligomer chain length). In all cases where n ≥ 1, [θ]333 increases linearly with increasing r between 0 and 0.32, and is accompanied by a corresponding decrease in [θ]275, which becomes negative as r approaches .32. Extrinsic band intensities reveal a dependence on n up to n = 5, above which increases in nonspecific binding result in a reduction in normalized band intensities. Polymer X shows a strong preference for B‐form nucleic acids and induces maximum extrinsic CD signal intensities with A‐T homopolymers. Alterations in helix hydration are believed to accompany complex formation. Inversions in [θ]275 of the octamer X‐poly(dA‐dT) complex have been attributed to the “alternating B” conformation of poly(dA‐dT).3 Similar inversions are not observed in other nucleic acid‐octamer X complexes. Visible and CD spectrometry data from competition studies in the presence of the antibiotics actinomycin D (AMD), daunomycin (DM), and distamycin A (DST) are consistent with “nonclassical” intercalation as the mode of binding, and these data place the potential binding site in or near the hydrophobic region of the minor groove. Reductions in [θ]333 with increasing urea further implicate the involvement of hydrophobic interactions in the formation of an asymmetric complex. Stabilization of the helix results in all cases as evidenced by alterations in Tm; corresponding changes, however, in cooperativity are not clearly discernable. Viscosity and light‐scattering data indicate no changes in molecular weight due to aggregation, and as such are not consistent with a transition to the ψ‐DNA upon complex formation.
Fluorescence decay of ethidium bromide in the presence of the Z‐conformation of poly‐(dG‐dC) and of poly(dG‐dC) modified by chlorodiethylenetriamine platinum(II) chloride
Authors: Genest D. , Malfoy B.
Published: December 2004

A time‐resolved fluorescence study of ethidium bromide (EB) in the presence of poly(dG‐dC) and of poly(dG‐dC) modified by chlorodiethylenetriamine platinum(II) chloride is presented under solvent conditions in which these polymers adopt the Z‐conformation (high ionic strength). It is shown that these polynucleotides can intercalate a very small quantity of EB. The binding parameters have been determined. The fluorescence lifetime of EB is slightly higher when bound to the Z‐conformation (≃25 ns) than when bound to the B‐conformation (≃23.7 ns). The nature of the salt has been checked. In the presence of 2.5M NaClO4, no transition from the Z‐conformation to another conformation is observed when EB is added. On the contrary, in the presence of 4.25M NaCl, EB induces a cooperative transition from the Z‐conformation to a conformation characterized by a much higher affinity for EB intercalation. In the case of poly(dG‐dC) this last conformation is identical to the one observed at low ionic strength (B‐conformation), but in the case of the platinated polymer this conformation is slightly different, as judged by the smaller value of the fluorescence lifetime of the intercalated EB.
Structural studies of poly(α‐aminoisobutyric acid)
Authors: Malcolm B. R. , Walkinshaw M. D.
Published: December 2004

The electron‐diffraction pattern of an oriented film of poly(α‐aminoisobutyric acid) in the 310‐helical conformation has been analyzed. The conformation was obtained by a linked‐atom least‐squares refinement of average values from crystal structures. Specimens treated with dichloracetic acid, to improve their crystallinity, conform to space group R3c with a = 21.8 Å, c = 5.95 Å. The structure contains channels that can accommodate molecules of dichloracetic acid. One molecule of acid per six residues fills the channels, and the R‐factor then is 34% using 23 reflections. Ir evidence is presented to show that the acid may hydrogen bond to the peptide groups. Some reflections occasionally observed on the diffraction photographs are attributed to a 15/4 α‐helix. The significance of the results is considered in relation to Aib‐containing peptides.
The relationship of bound water to the IR amide I bandwidth of albumin
Authors: Jakobsen R. J. , Wasacz F. M. , Brasch J. W. , Smith K. B.
Published: December 2004

Four different types of ir experiments, involving changes in pH, changes in pressure, and the use of nonaqueous solvents, and with either albumin molecules dissolved in saline or adsorbed albumin films, support the hypothesis that the bandwidth of the amide I vibration of albumin is directly related to the amount of bound water in this protein. From the amide I band narrowing and the amide I shift to higher frequencies, it is proposed that a more ordered helix structure results as the amount of bound water is decreased.
Adsorption of thionine on heparin studied by combining ultrafiltration and difference VIS spectroscopy
Authors: Nothelfer R. , Ruprecht J. , Heyl  , Baumgärtel H.
Published: December 2004

The combination of ultrafiltration and difference spectroscopy allows the quantitative determination of spectra of thionine bound to heparin. The spectra of the bound dye do not depend on the degree of coverage; this and the shape of the Scatchard plot show that “all‐or‐none” binding is valid. A calculus of variations based on a modification of the Hill plot shows that aggregates of seven thionine cations are bound. Tetrasaccharides with an average charge of two carboxylate and five sulfate groups are suggested to be the binding sites. The binding constant given for one mole thionine is 4.4 · 105 M−1. The Gibbs enthalpy for binding of one mole of thionine is −31.7 kJ·M−1 at 20°C.
Distribution of Mn2+ ions around poly(rI)·poly(rC)
Authors: Chang Ding‐Kwo , Kearns David R.
Published: December 2004

The distribution of bound Mn2+ ions about poly(rI)·poly(rC) has been studied by measuring the effect of this paramagnetic metal ion on the relaxation behavior of poly(rI)·poly(rC) protons. By combining selective spin – lattice and spin – spin relaxation rates for various protons, some of the principle regions of ion association can be identified. The relaxation data on the CH6 proton are consistent with a < 10% occupancy of phosphate inner‐sphere binding sites. The broadening of the imino proton resonance requires a substantial occupancy of sites located in the major groove, possibly near IN7. This would also be consistent with the observation that IH8 resonance is the proton most susceptible to relaxation by Mn2+. The relaxation data for the IH2 proton indicate a relatively low occupancy of minor‐groove binding sites (e.g., IN3).
Solution conformations of B. subtilis ribosomal 5S RNA: A calorimetric study
Authors: Chang Lee‐Hong , Marshall Alan G.
Published: December 2004

The unfolding of B. subtilis 5S RNA is examined by direct calorimetric measurement in the presence of various concentrations of Na+ and Mg2+. The composite differential scanning calorimetry (DSC) curve is analyzed into 3–5 individual two‐state melting transitions. In the absence of added Na+ or Mg2+, the 5S RNA segments melt together at Tm = 40°C. Addition of Na+ stabilizes the molecular structure (Tm = 56°C) and widens the melting temperature range, so that up to five component transitions are observed. Addition of Mg2+ alone produces a very stable structure (Tm = 75°C) with highly cooperative melting. Finally, addition of both Na+ and Mg2+ produces the highest stability (Tm = 76°C). The results are interpreted according to hypothetical secondary and tertiary base‐pairing schemes. The conformational changes demonstrated here may facilitate the movement of the protein synthesis machinery during RNA translation.
Interaction of fibrinogen‐binding tetrapeptides with fibrin oligomers and fine fibrin clots
Authors: Schindlauer Günther , Bale Marsha D. , Ferry John D.
Published: December 2004

The polymerization of fibrin, at pH 8.5 and ionic strength 0.45, and under conditions where the action of thrombin on fibrinogen was the rate‐determining step, was interrupted by inactivating thrombin with p‐nitrophenyl‐p′‐guanidinobenzoate (NPGB). Addition of the tetrapeptide Gly‐Pro‐Arg‐Pro (GPRP) partially dissociated the fibrin oligomers as shown by subsequent ligation with Factor XIIIa and calcium ion followed by denaturation and gel electrophoresis; polyacrylamide gel electrophoresis with reduction showed a decrease in the proportion of γ‐γ ligation compared with controls untreated by GPRP, and agarose gel electrophoresis showed a shift in the distribution of oligomer sizes. The dissociation was accomplished within 15 min and its extent was consistent with establishment of an equilibrium in which two molecules of GPRP react to sever an oligomer. When GPRP was introduced into fine unligated fibrin clots by diffusion, there was some dissociation as shown by differences in the degree of γ‐γ ligation after treatment by Factor XIIIa; but the action of GPRP was much slower and less complete than on soluble oligomers. However, even a small amount of dissociation affected the mechanical properties of fine clots profoundly. The shear modulus (measured 25 s after application of stress) decreased progressively with increasing concentration of GPRP introduced by diffusion. The rate of shear creep under constant stress and the proportion of irrecoverable deformation also increased enormously. If the steadystate creep rate is interpreted in terms of an effective viscosity, the latter is decreased by up to three orders of magnitude by the presence of GPRP. In terms of transient network theories of viscoelasticity, the average lifetime of a network strand is greatly diminished. However, the total density of strands remains constant during creep and creep recovery as shown by constancy of the differential modulus or compliance. Removal of GPRP by diffusion only partially restores the original shear modulus and creep behavior of the original clot. Some limited data on the effect of the tetrapeptide Gly‐His‐Arg‐Pro are also reported.