bluuebunny/crossref_metadata_2025 huggingface dataset:
Biophysical studies on fragments of the α‐factor receptor protein
Authors: Reddy A. P. , Tallon M. A. , Becker J. M. , Naider F.
Published: December 2004

The receptor for the α‐factor mating pheromone of the yeast Saccharomyces cerevisiae consists of 431 amino acid residues and is a member of a family of membrane proteins predicted to have seven transmembrane helices. Fragments of the receptor corresponding to two of the transmembrane helices [residues 246–269 (M6) and 273–302 (M7)], two of the interhelical loops [residues 107–125 (E2) and 191–206 (E3)], and to a portion of the carboxyl terminus [residues 350–372 (CT)] were synthesized using solid‐phase methodologies and purified to near homogeneity. CD was used to characterize the secondary structure of these peptides in trifluoroethanol (TFE), in TFE/water mixtures, in sodium dodecyl sulfate (SDS), and in the presence of dimyristoyl phosphatidylcholine (DMPC) liposomes. In TFE, M6 and M7 exhibited CD spectra consistent with highly helical peptides, whereas CT was partially helical. In contrast, E2 and E3 were either disordered or aggregated in this solvent. M6 did not partition well into DMPC vesicles whereas M7 remained helical. Both M6 and M7 assumed helical conformations in 25 mM SDS. The loop neptides and the carboxyl terminus peptide were either in a β‐structure or disordered in the presence of lipid. These findings represent the first biophysical evidence for conformations assumed by specific segments of the STE2 receptor protein. © 1994 John Wiley & Sons, Inc.
Computer simulation of the free energy of peptides with the local states method: Analogues of gonadotropin releasing hormone in the random coil and stable states
Authors: Meirovitch Hagai , Koerber Steven C. , Rivier Jean E. , Hagler Arnold T.
Published: December 2004

The Helmholtz free energy F (rather than the energy) is the correct criterion for stability; therefore, calculation of F is important for peptides and proteins that can populate a large number of metastable states. The local states (LS) method proposed by H. Meirovitch [(1977) Chemical Physics Letters, Vol. 45, p. 389] enables one to obtain upper and lower bounds of the conformational free energy, FB (b, l) and FA (b, l), respectively, from molecular dynamics (MD) or Monte Carlo samples. The correlation parameter b is the number of consecutive dihedral or valence angles along the chain that are taken into account explicitly. The continuum angles are approximated by a discretization parameter l; the larger are b and l, the better the approximations; while FA can be estimated efficiently, it is more difficult to estimate FB. The method is further developed here by applying it to MD trajectories of a relatively large molecule (188 atoms), the potent “Asp4‐Dpr10” antagonist [cyclo(4/10)‐(Ac‐Δ3Pro1‐D‐pFPhe2‐D‐Trp3‐Asp4‐Tyr‐5‐D‐Nal6‐Leu7‐Arg8‐Pro9‐Dpr10‐NH2)] of gonadotropin releasing hormone (GnRH). The molecule was simulated in vacuo at T = 300 K in two conformational states, previously investigated [J. Rizo et al. Journal of the American Chemical Society, (1992) Vol. 114, p. 2860], which differ by the orientation of the N‐terminal tail, above (tail up, TU) and below (tail down. TD) the cyclic heptapeptide ring. As in previous applications of the LS method, we have found the following: (1) While FA is a crude approximation for the correct F, results for the difference, ΔFA = FA(TD) – FA(TU) converge rapidly to 5.6(1) kcal/mole as the approximation is improved (i.e., as b and l are increased), which suggests that this is the correct value for ΔF; therefore TD is more stable than TU. (The corresponding difference in entrophy. TΔSA = 1.3(2) kcal/mole, is equal to the value obtained by the harmonic approximation.) (2) The lowest approximation, which has the minimal number of local states, i.e., based on b = 0 (no correlations) and l = 1 (the angle values are distributed homogeneously), also leads to the correct value of ΔF, within the error bars. This is important since the lowest approximation can be applied even to large proteins. (3) The method enables one to define the entropy of a part of the molecule and thus to measure the flexibility of this part. We have verified that the results for T[SA(TU) – SA(TD)] of the tail alone converged to 2.4(1) kcal/mole, which demonstrates the relatively high flexibility of the tail in the TU state. In order to study the random coil state, the Asp4‐Dpr10 analogue and its linear version were simulated by MU at 1000 K. We have been able to calculate a lower bound, ∼ 25 kcal/mole for T[S(linear) – S(cyclic)], which is the reduction in the conformational entropy caused by the ring closure. © 1994 John Wiley & Sons, Inc.
Ir and Raman spectra of L‐aspartic acid and isotopic derivatives
Authors: Navarrete J. T. López , Hernández V. , Ramírez F. J.
Published: December 2004

The ir and Raman spectra of L‐aspartic acid, L‐aspartic‐d4 acid, and L‐aspartic‐15N acid as solid samples have been recorded and measured. The data obtained were used to propose a general assignment of the fundamental vibrational modes in the basis of the isotopic shifts measured and correlations with previous data reported for other amino acids and related molecules. A MNDO‐scaled semiempirical force field was obtained, and vibrational frequencies and descriptions were used to support the previous assignments based on experimental features. © 1994 John Wiley & Sons, Inc.
Molecular dynamics simulation and nmr study of a blood group H trisaccharide
Authors: Widmalm Göran , Venable Richard M.
Published: December 2004

Molecular dynamics simulations in vacuum and solution have been carried out on 2′‐α‐L‐fucosyllactitol, a model for blood group H in conjunction with two‐dimensional nmr measurements on the same compound. Three independent starting conformations for the dynamics were chosen from low energy conformations obtained by a ϕ/ψ grid search. Nine 5 ns vacuum simulations of the trisaccharide were performed, employing three different ways to treat electrostatic interactions for each starting conformation: distance‐dependent dielectric with ε = r, constant dielectric with ε = 1, or constant dielectric with ε = 80. In vacuum, transitions of ϕ and ψ for the α‐L‐Fuc‐(1 → 2)‐β‐D‐Gal element occur in a cooperative manner. The virtual distance obtained for H1 in fucose to H2 in galactose from nuclear Overhauser effect spectroscopy experiments agree with one of the conformations of the trisaccharide in one of the three 100 ps aqueous simulations (ϕ/ψ ca. −100°/150°), indicating this may be a dominant solution conformation. The rms fluctuations of the ϕ‐ and ψ‐dihedral angles were ∼ 10° for a conformational state, both in the vacuum and the aqueous simulations. For the simulations in vacuum, the agreement with experimental NOE data is reasonable when a constant dielectric of 1 is used (major conformers having ϕ/ψ ca. −100°/150° and −140°/100°), whereas the agreement was poor with a constant dielectric of 80. Translational diffusion coefficients calculated from the simulation of the oligosaccharides were 0.12–0.18 × 10−5 cm2/s and from nmr measurements 0.27 × 10−5 cm2/s. © 1994 John Wiley & Sons, Inc.
Solution conformation of a cyclic neurokinin antagonist: A nmr and molecular dynamics study
Authors: Zhang Min , Quinn Thomas P. , Wong Tuck C.
Published: December 2004

The solution structure of a hexapeptide, cyclo(Gln‐Trp‐Phe‐Gly‐Leu‐Met), which is a selective NK‐2 antagonist, has been studied by a combination of two‐dimensional nmr and molecular dynamics (MD) techniques. The simulation based on nmr and MD data resulted in the convergence to a family of structures. Free molecular dynamics for 50 ps in the presence of DMSO solvent molecules shows that the structure is energetically stable. One intramolecular hydrogen bond between the amide proton of Gin and the carbonyl oxygen of Gly was revealed. This result is consistent with the results from the measurement of the temperature coefficient of the amide protons. The extent of intermolecular hydrogen bonding between the amide protons of the peptide and DMSO was also revealed by the free MD simulation. The resulting structure of the cyclic peptide contains a variation type I′ β‐turn in the Gly‐Leu‐Met‐Gln segment. Comparison of the structure of this peptide with that of other NK‐2 antagonist cyclic hexapeptides was made, and the activity of cyclic antagonists appears to be inversely related to the conformational rigidity of the cyclic peptides. © 1994 John Wiley & Sons, Inc.
Detection of monodisperse aggregates of a recombinant amelogenin by dynamic light scattering
Authors: Moradian‐Oldak J. , Simmer J. P. , Lau E. C. , Sarte P. E. , Slavkin H. C. , Fincham A. G.
Published: December 2004

Recombinant murine amelogenins M179 and M166 were expressed in Escherichia coli and purified. The aggregation properties of these amelogenins have been investigated in aqueous solutions as well as acetonitrile‐containing solutions using dynamic light scattering. Dynamic light scattering provides direct measurement of the translational diffusion coefficient and hydrodynamic radius, and of an estimate of the molecular weight. Polydispersity and statistical parameters of how to interpret the analysis are also provided. Amelogenin aggregation was examined in solutions of a range of pH, ionic strengths, and protein concentrations. It was shown that at pH 7.8–8 and ionic strength of 0.02–0.05M the M179 molecules form monodispersed aggregates with hydrodynamic radii ranging from 15 to 19 nm. Analysis of hydrodynamic radii and size distribution of M179 aggregates in acetonitrile‐containing solvents compared to that in aqueous solutions indicated a primary role for hydrophobic interactions in the association process of amelogenin molecules to form aggregates. Comparison between the aggregates formed by M179 and M166, which lacks the hydrophilic carboxy‐terminal 13 residue sequence of M179, suggested that the self‐assembly of amelogenin molecules to form stable and monodisperse aggregates requires the presence of the hydrophilic carboxy‐terminal sequence of M179. © 1994 John Wiley & Sons, Inc.
Defect peptide chemistry: Perturbations in the structure of a homopentapeptide induced by a guest residue interrupting side‐chain regularity
Authors: Benedetti Ettore , Pedone Carlo , Pavone Vincenzo , Di Blasio Benedetto , Saviano Michele , Fattorusso Roberto , Crisma Marco , Formaggio Fernando , Bonora Gian Maria , Toniolo Claudio , Kaczmarek Krzysztof , Redlinski Adam S. , Leplawy Miroslaw T.
Published: December 2004

The fully blocked pentapeptide Tfa‐(Deg)2‐L‐Abu‐(Deg)2‐OtBu (Tfa:triflouroacetyl; Deg: Cα,α‐diethylglycine; OtBu: tert‐butoxy) adopts in the crystal state a regular, right‐handed 310‐helical structure stabilized by three N  H …︁ O  C intramolecular 1 ← 4 (or C10) H bonds, as determined by an x‐ray diffraction analysis. However, a Fourier transform ir absorption and 1H‐nmr study strongly supports the view that in deuterochloroform solution the four Deg residues at both termini of the peptide main chain are involved in successive, fully extended C5 forms. A comparison with the stable, fully developed, multiple C5 conformation of Tfa‐(Deg)5‐OtBu indicates that incorporation of an Abu guest residue, interrupting the side‐chain uniformity of the host (Deg)5 homopeptide, while altering only marginally the conformation in a solvent of low polarity, is responsible for a dramatic perturbation of the crystal‐state structure. © 1994 John Wiley & Sons, Inc.
A conformational study of agarose by vacuum uv CD
Authors: Arndt Edward R. , Stevens Eugene S.
Published: December 2004

Agarose films were prepared using a variety of casting conditions, heat treatments, and solvation treatments. Films represented the high temperature sol, the room temperature gel, and a high temperature state with cross‐links intact. The vacuum uv CD of the film samples varies considerably, but the similarities between the film CD and the previously reported solution CD show that dehydration per se does not affect the CD. Combining the CD results for the dried sol with the x‐ray results of Foord and Atkins leads to a description of the high temperature sol in terms of locally extended chain conformations. The CD of the gel and its peculiarly high uv absorption are satisfactorily, albeit not uniquely, interpreted in terms of the double‐helix model of the gel, in agreement with previous conformational studies based on chiroptical properties. © 1994 John Wiley & Sons, Inc.
Helix formation in model peptides based on nucleolin TPAKK motifs
Authors: Xu Xiaobing , Cooper Laura G. , DiMario Patrick J. , Nelson Jeffrey W.
Published: December 2004

The structures formed by peptide models of the N‐terminal domain of the nucleolar protein nucleolin were studied by CD and nmr. The sequences of the peptides are based on the putative nucleic acid binding sequence motif TPAKK: The peptides TP1 and TP2 have the sequence acetyl‐G(ATPAKKAA)nG‐amide, with n = 1 and 2, respectively. CD measurements indicate structural changes in both peptides when the lysine side chains are uncharged by increasing the pH or acetylation of the side‐chain amines. When trifluoroethanol (TFE) is added, more extensive structural changes are observed, resembling helical structure based on nmr nuclear Overhauser effect (NOE) and Cα proton chemical shift changes, and CD spectra. The structure formed in 0.5M NaClO4 as observed by nmr is similar to that when the lysine side chains are acetylated, due presumably to interactions of perchlorate ion with side‐chain charges on lysines. The helical structure observed in TPAKK motifs may be stabilized via N‐capping interactions involving threonine. The structures observed in TFE suggest that the Thr‐Pro sequence initiates short helical segments in TPAKK motifs, and these helical structures might interact with nucleic acids, presumably via interactions between lysines and threonines of nucleolin. © 1995 John Wiley & Sons, Inc.
Normal mode calculation of a netropsin–DNA complex: Effect of structural deformation on vibrational spectrum
Authors: Chen Y. Z. , Prohofsky E. W.
Published: December 2004

Calculations of the normal mode spectrum of a netropsin‐DNA complex, an isolated DNA helix of the same sequence, and a free netropsin molecule were carried out and compared with observations. We find that the frequency of modes of the complex that are primarily vibrations localized to the helix are relatively unchanged from the frequencies found for similar modes of the isolated helix. On the other hand, the frequencies of those modes of the complex that are found to be primarily in the netropsin are found to change significantly in the complex from that of the free netropsin. Further analysis indicates that the changes in the “netropsin modes” are primarily due to the conformational deformation induced in the netropsin by formation of the complex. All the frequencies and shifts in frequency are in good agreement with recent Raman measurements. © 1995 John Wiley & Sons, Inc.
Molecular conformational analyses of dehydroalanine analogues
Authors: Alemán Carlos , Casanovas Jordi
Published: December 2004

The conformational preferences of dehydroalanine (ΔAla) were examined through ab initio calculations. The geometries of the minimum energy conformations for N‐formyldehydro alanilamide and N‐acetyl‐N′‐methylamide of dehydroalanine were determined by gradient optimization at the HF/6‐31G* level, and correlation corrections were examined with MP2 single‐point energy calculations. Furthermore, HF/3‐21G ab initio geometry optimizations were performed on nine conformations of the model tripeptide N‐acetyl‐N′‐methylamide of didehydroalanine. The results indicate that the C5 is the lowest energy conformation at all levels of theory. However, the relative energy of the helix conformation decreases when the number of ΔAla residues in the peptide chain increases. On the other hand, significant variations of the geometry upon conformational change were observed for the three compounds investigated. These results permit to extract important conformationally dependent geometry trends. The results of this study were compared to x‐ray diffraction data on single crystals of dehydroalanine‐containing peptides. © 1995 John Wiley & Sons, Inc.
Conformational analysis of endothelin‐1: Effects of solvation free energy
Authors: Hempel J. C. , Fine R. M. , Hassan M. , Ghoul W. , Guaragna A. , Koerber S. C. , Li Z. , Hagler A. T.
Published: December 2004

In order to investigate conformational preferences of the 21‐residue peptide hormone endothelin‐1 (ET‐1), an extensive conformational search was carried out in vacuo using a combination of high temperature molecular dynamics / annealing and a Monte Carlo / minimization search in torsion angle space. Fully minimized conformations from the search were grouped into families using a clustering technique based on rms fitting over the Cartesian coordinates of the atoms of the peptide backbone of the ring region. A wide range of local energy minima were identified even though two disulfide bridges (Cys1‐Cys15 and Cys3‐Cys11) constrain the structure of the peptide. Low energy conformers of ET‐1 as a nonionized species in vacuo arestabilized by intramolecular interaction of the ring region (residues 1‐15) with the tail (residues 16–21). Strained conformations for individual residues are observed. Conformational similarity to protein loops is established by matching to protein crystal structures

In order to assess the influence of aqueous environment on conformational preference, the electrostatic contribution to the solvation energy was calculated for ET‐1 as a fully ionized species (Asp8, Lys9, Glu10, Asp18, N‐ and C‐terminus) using a continuum electrostatics model (DelPhi) for each of the conformed generated in vacuo, and the total solvation free energy was estimated by adding a hydrophobic contribution proportional to solvent accessible surface area. Solvation dramatically alters the relative energetics of ET‐1 conformers from that calculated in vacuo. Conformers of ET‐1 favored by the electrostatic salvation energy in water include conformers with helical secondary structure in the region of residues 9–15.

Perhaps of most importance, it was demonstrated that the contribution tosolvation by an individual charge depends not only on its solvent accessibility but on the proximity of other charges, i.e., it is a cooperative effect. This was shown by the calculation of electrostatic solvation energy as afunction of conformation with individual charges systematically turned “on” and “off”. The cooperative effect of multiple charges on solvation demonstrated in this manner calls into question models that relate solvation energysimply to solvent accessibility by atom or residue alone. © 1995 John Wiley & Sons, Inc.
Comparison of base inclination in ribo‐GC and deoxybribo‐GC polymers, and synthesis of poly(rGrC)‐poly(rGrC)
Authors: Jin Xiaokui , Johnson W. Curtis
Published: December 2004

The inclination angle between the base normal and the helix axis, and the axes around which the bases incline, are measured for ribo‐GC polymers in buffer by using flow linear dichroism (LD), and compared to measurements for deoxyribo‐GC polymers in buffer and under dehydrating conditions. A new method is designed to synthesize poly(rGrC) ‐poly(rGrC), which is not available commercially, in large quantities. The LD of this RNA reveals inclination angles that are similar to the B‐form DNA in buffer, although the axes are different. The CD of poly(dGdC)‐poly(dGdC) under the dehydrating conditions is similar to poly(rGrC)‐poly(rGrC), indicating it is in the A form, and the LD gives larger inclination angles than either the B form or the corresponding RNA. Poly(dG)‐poly(dC) is in the A form in buffer. Comparison among poly(rG)‐poly(rC) in buffer, and poly (dG)‐poly(dC) in buffer and under dehydrating conditions, reveals similar inclination angles and axes, although the LD shows that the DNA has the largest inclination angles. Except for poly(rGrC)‐poly(rGrC), which has a unique reduced dichroism, all the axes for G are similar, as are the axes for C. © 1995 John Wiley & Sons, Inc.
The use of spectral decomposition via the convex constraint algorithm in interpreting the CD‐observed unfolding transitions of C coils
Authors: Holtzer Marilyn Emerson , Holtzer Alfred
Published: December 2004

Decomposition of CD spectra for the unfolding of both coiled‐coil and single‐helical molecules is carried out via the convex constraint algorithm (CCA) [A. Perczel, M. Hollósi, G. Tusnády, and G. D. Fasman (1991) Protein Engineering, Vol. 4, pp. 669–679]. Examined are (1) our thermal unfolding data for rabbit αα‐tropomyosin and chicken gizzard γγ‐ tropomyosin coiled coils, and for a35‐residue, tropomyosin‐model peptide that forms single helices, not coiled coils; (2) extent pH‐induced unfolding data for 50‐ and 400‐residue poly‐L‐glutamic acid. Each set of spectra shows a sharp isodichroic point near 203 nm. We find here that the CCA is of sharply limited use for analyzing such data. The component spectra obtained for a given substance not only depend on the particular experimental spectra included and on the chosen number of component spectra, but all pass through the experimental isodichroic point. The latter is physically unlikely for more than three component spectra, and physically impossible for conformers, such as β structures, having known isodichroic points elsewhere. Our conclusions are in contrast to those of an extant decomposition via CCA of thermal spectra for rabbit αα‐tropomyosin [N. J. Greenfield and S. E. Hitchcock‐DeGregori(1993) Protein Science, Vol. 2, pp. 1263‐1273] that postulates the existence of five conformers, including β structures, in the unfolding. Moreover, an extant diagnostic based on the θ222/θ208 ratio and allegedly distinguishing between spectra for coiled coil and for single α‐helix is shown here to be unreliable. © 1995 John Wiley & Sons, Inc.
Hydration of cis and trans N‐methylformamide as revealed by the use of 17O‐NMR, molecular mechanics, and ab initio calculations
Authors: Gerothanassis Ioannis P. , Demetropoulos Ioannis N. , Vakka Constantina
Published: December 2004

The solvation of cis and trans N‐methylformamide (NMF) by water was investigated using a combination of 17O‐nmr spectroscopy, classical molecular mechanics [MM2(77) and MM2(87)] force field, and ab initio 4‐31G* gradient optimization calculations. In dilute aqueous solution, the 17O‐nmr spectra of NMF indicate strong shielding by 66.9 and 66.1 ppm for the cis and trans amide oxygens, respectively, compared to those values obtained in dilute toluene solution. This demonstrates that both isomers are equally solvated by molecules of water, which are further hydrogen bonded to molecules of water of the bulk solvent. Molecular mechanics simulations were carried out for cis and trans NMF in a cluster of water molecules. Radial distribution functions show structural contacts by several water molecules at the amide CO and NH group, which are significantly more pronounced with MM2 (87) calculations. Ab initio 4‐31G* gradient optimization calculations on the supermolecule trans NMF‐(H2O)3 indicates the presence of more than two hydrogen‐bond contacts at the carbonyl oxygen. This is in agreement with MM2 calculations and provides further evidence for multiple acceptor properties of the amide oxygen and an out of the amide plane arrangement of the bound molecules of water. Comparison of the integration data to the first radial distribution function (rdf) minima shows that the local solvation of the CO and NH groups is very similar for both cis and trans isomers. The intermolecular geometric parameters of the supermolecule trans NMA–(H2O)3 and the first rdf maxima resulting from MM2 (87) and MM2 (77) calculations are compared with distribution of water molecules around the CO and NH groups of peptides and proteins resulting from x‐ray and neutron diffraction experiments. The rdfs involving the methyl group of NMF demonstrate the nonrandom distribution of solvent sites with first maxima in reasonable agreement with distribution of water molecules around the apolar side chain of amino acid residues in proteins. © 1995 John Wiley & Sons, Inc.
Influence of proline‐14 substitution on the secondary structure in a synthetic analogue of alamethicin
Authors: Brachais Laurent , Duclohier Hervé , Mayer Catherine , Davoust Daniel , Molle Gérard
Published: December 2004

Due to the bend introduced by proline 14 in the conformation of alamethicin (AcUPUAUQUVUGLUPVUUEQFol), the role of this residue was assumed essential in the barrel‐stave model for voltage‐gated ion channels. Taking advantage of a previous synthetic alamethicin analogue (L2) in which all eight α‐aminoisobutyric(U) Were replaced by leucines (AcLPLALAQLVLGLLPVLLEQFol), another analogue (L5) was synthesized in order to test the effects ofproline‐14 substitution by an alanine (AcLPLALAQLVLGLLPVLLEQFol). Previous conductance experiments showed that both high voltage dependence and multistate behavior were conserved. In order to complement these functional results, a conformational study of L5 has been undertaken and compared to L2 using CD. high field nmr, and molecular dynamics. Results show that L5 presents a better ordered structure than L2 particularly in the region of the substitution and in the C‐terminal part. These results are discussed as regards the previous hypothesis of the nonessential character of helix bending for the gating of voltage‐dependent ion channels. © 1995 John Wiley & Sons, Inc.
Properties of auricularia auricula‐judae β‐D‐glucan in dilute solution
Authors: Zhang Lina , Yang Liqun
Published: December 2004

A Water‐soluble glucan A was isolated from the fruit body of Auricularia auricula‐judae. It is composed of a backbone chain of β‐(1 → 3)‐linked D‐glucose residues, two out of three glucose residues being substituted at C‐6 positions with a single glucose unit. The weight average molecular weight Mw, number average molecular weight Mn, and intrinsic viscosity [η] of the fractionated samples were studied at 25°C in water and in dimethylsulfoxide (DMSO). The Mark‐Houwink equation was established as [η] = 6. 10 × 10−4 Mw1.14 for the glucan A having Mw ranging from 9 × 105 to 1.6 × 106 in water. The values of [η] in water are far higher than those in DMSO, but the values of Mn measured in water are the same as those in DMSO. Analysis of Mw and [η] in terms of the known theories for rods and wormlike chains yielded 1030 ± 100 nm−1, 90 ± 20 nm, 1.3 ± 0.3 nm, and 0.26 ± 0.03 nm for molar mass per unit contour length ML, persistence length q, diameter d, and contour length h per main‐chain glucose residue, respectively. The present data suggest that glucan A dissolves in water as single‐stranded helical chains and in DMSO as semiflexible chains. © 1995 John Wiley & Sons, Inc.
Anomalous temperature fluorescence quenching of N‐Trp terminal peptides
Authors: Brancaleon L , Crippa P. R. , Diemmi D
Published: December 2004

The photophysics of Trp‐containing peptides is extremely affected by the position of the indole ring with respect to the substituents. In this work an unusual temperature fluorescence quenching behavior is presented. The N‐tryptophan terminal peptides (N‐ Trp) show an increase of the static emission intensity as rising the temperature from 10 to about 40°C. The anomaly is typical of the N‐Trp terminal peptides since neither tryptophan (Trp) nor glycyl‐tryptophan (Gly‐Trp) and alanyl‐tryptophan (Ala‐Trp) show the same trend; a similar behavior is not detected in the C‐tryptophan terminals. The other important features are the wavelength and pH dependence of the effect. The anomaly is in fact detected only at neutral pH and for excitation wavelength near the red edge of the UVB absorption band of indole. An interpretation of the anomaly is suggested, though more sophisticated techniques are needed to better focus the problem; the model proposed involves the superimposition of a ground state effect (the temperature‐induced equilibrium shift from the zwitterionic to the anionic form of the peptides) and an excited state mechanism. At present no unique interpretation can be provided about the excited state mechanism that favors the anomaly and some suggestions are discussed. © 1995 John Wiley & Sons, Inc.
Novel biopolymers for drug discovery
Authors: Moran Edmund J. , Wilson Troy E. , Cho Charles Y. , Cherry Sara R. , Schultz Peter G.
Published: December 2004

The solid phase synthesis and generation of libraries of “unnatural biopolymers” is described. These polymers are characterized by novel backbones and building blocks, the properties of which may modify their pharmacological and folding properties. © 1994 John Wiley & Sons, Inc.
Peptides in membranes: Helicity and hydrophobicity
Authors: Deber Charles M. , Li Shun‐Cheng
Published: December 2004

Synthetic model membrane‐interactive peptides—both of natural and designed sequence—have become convenient and systematic tools for determination of how the membrane‐spanning segments within integral membrane proteins confer protein structure and biology. Conformational studies on these peptides demonstrate that the α‐helix is the natural choice of conformation for a peptide segment in a membrane, and that a helical conformation will arise “automatically” in a peptide above a threshold hydrophobicity that allows it to associate stably with the membrane. Environmental and sequential contexts thus impart conformational versatility to many of the amino acids, thereby providing a mechanism for producing the diverse structural and functional properties of proteins. © 1994 John Wiley & Sons, Inc.
The suitability of a monofunctional reagent of an undecagold cluster for phasing data collected from the large ribosomal subunits from Bacillus stearothermophilus
Authors: Bartels Heike , Bennett William S. , Hansen Harly A. S. , Eisenstein Miriam , Weinstein Shulamith , Müssig Jutta , Volkmann Niels , Schlünzen Frank , Agmon Ilana , Franceschi Francois , Yonath Ada
Published: December 2004

An electron density map of the large ribosomal subunit from Bacillus stearothermophilus was obtained at 26 Å resolution by single isomorphous replacement (SIR) from a derivative formed by specific quantitative labeling with a dense undecagold cluster. For derivalization, a mono‐functional reagent of this cluster was bound to a sulfhydryl group of a purified ribosomal protein. which was in turn reconstituted with core particles of a mutant lacking this protein. The native, mutated, and derivatzed 50S ribosomal subunits crystallize under the same conditions in the same space group. Under favorable conditions, crystals of the derivatized subunit proved to be isomorphous with the native ones, whereas the crystals of the mutant may have somewhat different packing.

After resolving the SIR phase ambiguity by solvent flattening, the electron density shows a packing that is consistent with the noncrystallographic symmetry found by Patterson searches as well as with the motif observed in electron micrographs of thin sections of the crystals. These studies established that phase information can be obtained from heavy metal clusters, even when the crystals under investigation are unstable and weakly diffracting. These results encouraged further effort at the construction of specifically derivatized crystals from other ribosomal particles that diffract to higher resolution. © 1994 John Wiley & Sons, Inc.
Effect of UV‐irradiated nitrogen gas on biological nitrogen fixation in batch cultures of Azotobacter vinelandii
Authors: Zacharias Bengt
Published: December 2004

In batch cultures of Azotobacter vinelandii exposed to air and ultraviolet‐irradiated nitrogen, an increases in nitrogen fixation, compared to that of cultures exposed to air and non‐irradiated nitrogen, has been noted. This supports the hypothesis that an ionized nitrogen molecule is important for the biological nitrogen fixation.
Production of eburicoic acid by submerged cultures of Polyporus sulfureus
Authors: Pan S. C. , Frazier W. R.
Published: December 2004

A process has been developed whereby eburicoic acid, a triterpenoid acid, can be produced by growing Polyporus sulfureus in submerged form in either shake flasks or tank fermentors equipped with aeration and agitation systems.
Volume increase of kernels of corn and sorghum accompanying absorption of liquid water
Authors: Fan Liang‐Tseng , Chu Pu‐Shan , Shellenberger John A.
Published: December 2004

The linearity between weight and volume increases, which takes place when kernels of corn and sorghum are immersed in water, was established experimentally. The rate of volume increase was correlated according to an equation analogous to the diffusion equation. The coefficients of volume increase obeyed the Arrehenius equation. The results would be of practical value in designing steeping vessels, as well as of theoretical interest in elucidating the mechanism of water diffusion in cereal grains.
Experiments with lysis of living cells of Eremothecium ashbyii and of Methanomonas by microbial enzymes
Authors: Carenberg C. O. , Hedén C.‐G.
Published: December 2004

The possibility to use microorganisms as human food is limited by several factors. The intact cell is resistant to digestion, the cell wall is unbalanced in essential amino acids, and the nucleic acids are said to be harmful. For using single cell protein as food it may thus be necessary to disrupt the cell wall and separate the protein from nucleic acid. This paper is concerned with the production and properties of extracellular enzymes able to lyse cell walls of microorganisms. Soil bacteria and actinomycetes have been cultivated and lytic enzymes from these organisms have been used to lyse living cells of the yeast like organism E. ashbyii. Efforts were also made to use these enzymes for lysing cell of a Methanomonas sp.
Respiratory relationships of a symbiotic algal‐bacterial culture for wastewater nutrient removal
Authors: Humenik F. J. , Hanna G. P.
Published: December 2004

A continuous symbiotic algal‐bacterial system was developed consisting essentially of a mixed Chlorella‐activated sludge culture which would efficiently remove nutrients from wastewater under aerobic conditions without supplementary aeration. Oxygen decline data were fitted to a mathematical model used to predict respiratory rates, photosynthetic oxygenation, and steady‐state oxygen concentrations. Stable relative biological populations and a dissolved oxygen concentration of about 2 mg/1 were maintained during steady‐state operation with daily harvesting of excess biomass. Respiratory and physiological relationships indicated that the carbon dioxide‐oxygen balance is a primary control that governs the steady‐state operation of a symbiotic algal‐bacterial culture. The close association of the algae and bacteria resulted in an algal‐bacterial floc with settled rapidly yielding a clear supernatant.
Dialysis fermentation. II. Kinetic analysis of salicylic acid production via cell growth and maintenance
Authors: Abbott Bernard J. , Gerhardt Philipp
Published: December 2004

A kinetic analysis was made of the relationship between salicylate production from naphthalene and growth of Pseudomonas fluorescens in semicontinuous dialysis culture. The specific rates both of product formation and growth initially were increased by the diffusional withdrawal of salicylate, but subsequently were reduced to low levels despite continued salicylate removal. Productivity and growth were correlated by the Luedeking‐Piret equation in an initial nondialysis period and in the early stages of dialysis fermentation, when specific growth rates exceeded. 005 hr−1. Below this level of growth at later stages of dialysis fermentation, the specific production rate was correlated only with total cell mass by a proportionality constant of .035 hr−1, which was attributed to maintenance metabolism. Maintenance accounted for about 84% of the total salicylate produced, while growth‐associated metabolism accounted for the remainder.
The supply of oxygen to submerged cultures of BHK 21 cells
Authors: Radlett P. J. , Telling R. C. , Whitside J. P. , Maskell M. A.
Published: December 2004

Oxygen solution rates were measured in 4, 30, and 100 liter culture vessels, and the oxygen demand of growing BHK 21 cells estimated. This data was used to calculate the minimal sparged air rates necessary to satisfy oxygen demand throughout the cell growth cycle, and in this way adequate oxygen was supplied without the damaging effects of excessive sparging. Comparable results were obtained when oxygen was supplied by this method and when pO2 was controlled at 80 mmHg, but both cell growth rate and maximum cell density were reduced when pO2 was controlled at other values.
Growth models of cultures with two liquid phases. VII. Substrate dissolved in dispersed phase; effect of dispersed phase volume and temperature
Authors: Prokop A. , Erickson L. E.
Published: December 2004

The effects of dispersed phase volume and temperature on the batch growth of Candida lipolytica on gas oil are investigated. Growth parameters are presented for two sets of experiments. The shape of growth curves was basically similar to the system composed of n‐hexadecane dissolved in dewaxed gas oil, in spite of the complex nature of the substance. All of the batch growth curves exhibited a linear growth region. The rate of linear growth and its length varied with change in dispersed phase volume. The effect of temperature on growth rate was investigated for temperatures ranging from 23°C to 34°C. The results show a smaller activation energy during linear growth than during the early stages of batch growth. These results are analyzed from the viewpoint of growth models presented previously. The results indicate that growth at drop surfaces is important and that segregation effects may be important.
Microbiological studies on the formation of gramicidin S synthetases
Authors: Matteo C. C. , Glade M. , Tanaka A. , Piret J. , Demain A. L.
Published: December 2004

Rapid and extensive growth of Bacillus brevis ATCC 9999 was obtained in a complex medium containing yeast extract and peptone. Gramicidin S (GS) production in this medium reached 2.5 g/liter and 0.25 g/g dry cell weight. GS synthetase I production was also high in this complex medium. Chemically defined media were also developed for this strain. In a glycerol‐ammonium sulfate‐Tris‐salts medium, the culture grew about 40% as well (rate and extent) as in complex medium. Although GS production was low (0.23 g GS/liter), peak specific activity of GS synthetase I was as high as on complex medium. Nutritional experiments showed that growth was stimulated by glutamine, methionine, proline, arginine, and histidine. Addition of these amino acids almost doubled the rate and extent of growth and GS production on a volumetric basis. However the increase in GS was due merely to the increased cell density; GS synthetase I specific activity was in fact decreased by the supplement. Complex medium is better than defined medium for GS and GS synthetase production due to increased cell density and a slower rate of synthetase disappearance.
Solution of immobilized enzyme problems by collocation methods
Authors: Ramachandran P. A.
Published: December 2004

A method of solution to the problem of enzyme effectiveness factor has been presented. For a rapid estimation of the same, a graphical procedure is discussed which is sufficiently accurate for many practical situations. Applications to systems with rate dependent on position in the pellet and substrate and product inhibition is discussed. The case of concentration dependent diffusivity (facilitated diffusion) can also be solved by a simple transformation.
Microbial growth on hydrocarbons—some experimental results
Authors: Velankar S. K. , Barnett S. M. , Houston C. W. , Thompson A. R.
Published: December 2004

A simple mechanism of hydrocarbon uptake by microorganisms is examined. This model considers the presence of micelles of surface active agents as essential for growth of microbial cells on hydrocarbon substrates. Larger hydrocarbon droplets serving as reservoirs may also be present. Experimental results to support the model are presented. The results of others are explained on the basis of this model.
Behavior of subcellular marker enzymes during the Hela cell cycle
Authors: Thilly W. G. , Arkin D. I. , Nowak T. S. , Wogan G. N.
Published: December 2004

Six enzymes associated with the activities of the nucleus (thymidine kinase), mitochondria (succinate dehydrogenase), lysosomes (acid phosphatase), peroxisomes (catalase), (cytosol lactate dehydrogenase), and the intermembranal mitochondrial space (alkaline DNase) were assayed at 2 hr intervals over the division cycle of repetitively resynchronized HeLa cells. The results indicated a high degree of reproductibility for cells synchronized by the method of perpetual resynchronization and may be of direct use to those interested in subcellular organellogenesis.
Kinetics of a hollow fiber dehydrogenase reactor
Authors: Fink David J. , Rodwell. Victor W.
Published: December 2004

A hollow fiber module was used as a reactor for conversion of ethanol to acetaldehyde in the presence of horse liver alcohol dehydrogenase as catalyst. Mass transport rates for NAD+, the overall acetaldehyde generation rate, catalyst effectiveness factors, and the overall order of the reaction with respect to NAD+ concentration were measured. A coupled‐substrate reactor with continuous in situ regeneration of cofactor was also examined. Two substrates of opposite redox state were added simultaneously to the feed stream. NADH and acetaldehyde concentrations were monitored in the effluent stream. The cofactor recycle number, or ratio of moles of product to moles of NADH produced, exceeded 10,000 under certain conditions. While decreasing the NAD+ concentration in the feed stream decreased reactor productivity somewhat, it greatly enhanced the ratio of product formed per mole of NAD+ fed to the reactor. It is suggested that high cofactor costs in dehydrogenase reactors may be overcome with efficient in situ regeneration and secondary recovery and recycling of cofactor from the process stream.
Nitrate and nitrite reduction by liquid membrane‐encapsulated whole cells
Authors: Mohan Raam R. , Li Norman N.
Published: December 2004

Purified enzymes and cell‐free homogenates encapsulated by liquid‐surfactant membrane have been shown to retain their catalytic activity (see previously published literature). This paper describes the preparation and properties of liquid‐surfactant membrane‐encapsulated whole cells of Micrococcus denitrificansATCC 21909. Batch and continuous studies with this model system have demonstrated that encapsulated viable cells reduce nitrates and retain their catalytic activity over anextended period of time. In batch operation, the reactivity of the encapsulated whole cells has been investigated under a variety of experimental conditions. The system is capable of reducing NO3− or NO2−. Data obtained indicate that encapsulated live cells have a broad pH and temperature optimum range. The encapsulated cells remain viable and do not “escape” into the external aqueous phase, even after five days of constant stirring with nitrate‐containing simulated wastewater. Pulsed substrate addition experiments have demonstrated that the encapsulated cells also effectively reduce NO2− with no significant reduction in activity, even after 5.5 days of incubation at 30°C. The membrane selectivity for ion transfer has been achieved by incorporating oil‐soluble ion exchangers in the membrane. Because of the protection of the liquid membranes, the catalytic reduction of NO2− by the encapsulated whole cells is not inhibited by 1 × 10−4 M mercuric chloride, which is otherwise extremely toxic to the cells, when present in the external aqueous phase. Continuous reduction of 20 ppm of NO2− by liquid membrane‐encapsulated whole cells has been demonstrated in a constantly stirred reactor over a test period of about one week. In this paper we will discuss the reduction of NO3−and NO2− by the liquid membrane‐encapsulated whole cells of M. denitrificansATCC 21909 mainly in batch runs undera variety of experimental conditions, such as cell and substrate concentrations, product and inhibitor permeation, pH and temperature, effect of oil‐soluble ion exchangers on the substrate diffusion, etc.
Single‐cell protein production by photosynthetic bacteria cultivation in agricultural by‐products
Authors: Shipman R. H. , Kao I. C , Fan L. T.
Published: December 2004

The growth of the nonsulfur photosynthetic bacterium Rhodopseudomonas gelatinosa was investigated as a possible way to produce single‐cell protein from agricultural by‐products. Of the various raw materials examined as potential feedstocks, wheat bran infusion was selected as the substrate for mass culture and continuous cultivation studies. Harvested photosynthetic cells contained approximately 65.0% crude protein and 5.1% nucleic acid (RNA). The amino acid content of harvested photosynthetic proteins was comparable with conventional proteins of plant and animal origin.
The effect of methanol, formaldehyde, and formic acid on growth of Candida boidinii 11 Bh
Authors: Pilát P. , Prokop A.
Published: December 2004

The dependence of the specific growth rate of Candida boidinii 11 Bh on methanol concentration follows Monod‐type kinetics only in a narrow concentration range (0–0.8% v/v), with a saturation constant of about 19 mmol. With a further increase in methanol concentration of up to 3% v/v, the specific growth rate remains constant. Beyond this level, the growth rate gradually drops to zero, reaching the threshold value at 7.8% v/v. The specific growth rate is also strongly dependent on the size of the inoculum. The biomass yield decreases with an increase in the methanol concentration. Formaldehyde and formic acid, oxidative products of methanol, approximately 100 and ten times, respectively, more toxic to growth than the original substrate.
Comparison of batch and semicontinuous cultures for production of protein from mesquite wood by Brevibacterium sp. JM98A
Authors: Fu Tzulin Tom , Thayer D. W.
Published: December 2004

The production of protein by a Brevibacterium sp. JM98A usingmesquite wood as the substrate was compared in batch and semicontinuous cultures. A 14 liter glass fermentor with automatic pH, temperature, and foam control was used for the study. A pH range of 6.6 to 7.2 was optimum for the growth of JM98A. The batch and semicontinuous cultures were compared on the basis of viable cell counts, protein production, CMC‐Ase (β‐1,4‐glucanase) activity, and filter paper cellulase (β‐1,4‐glucan cellobiohydrolyase) activity. Total hexose, cellulose, and reducing sugar consumption were measured. The semicontinuous process yielded 2.97 times as much protein in 72 hr as the batch cultures. Most of the biomass resulted from the utilization of soluble sugars rather than from the degradation of cellulose during the semicontinuous process.
Response of a continuous anaerobic culture to periodic variation of the feeding mash concentration
Authors: Borzani Walter , Gregori Rosana E. , Vairo Marina L. R.
Published: December 2004

The authors studied the influence of periodic variations (one cycle per 24 hr) of the feeding mash concentration of a continuous anaerobic culture of Saccharomyces cerevisiae in sugarcane molasses media at constant dilution rate. It was observed that the average yield coefficients during the transient state were practically equal to the yield coefficient obtained during steady‐state experiments. It was also observed, in each experiment, that the average specific growth rate during the transient state was equal to the dilution rate.
Invertase biosynthesis by Saccharomyces carlsbergensis in batch and continuous cultures
Authors: Toda K.
Published: December 2004

The biosynthesis of invertase by Saccharomyces carlsbergensis LAM 1068 was studied in relation to its glucose effect at both unsteady and steady states of growth. Experimental correlations between the dilution rate and invertase specific activity (E/X) in chemostat, cultures led to an optimum for the enzyme synthesis at a particular intermediate growth rate. The value of E/X increased from 1.1 (U/mg biomass) in batch cultures to 13 (U/mg biomass) in chemostat cultures. A mutant strain A3 showed the highest value for E/X = 25 (U/mg biomass) at high dilution rates where glucose repression was observed with the wild strain.