X-Ray Studies On Crystalline Complexes Involving Amino-Acids And Peptides .11. Peptide Aggregation And Water-Structure In The Crystals Of A Hydrated 1-1 Complex Between L-Histidyl-L-Serine And Glycyl-L-Glutamic Acid

The hexahydrate of a 1:1 complex between L-histidyl-L-serine and glycyl-L-glutamic acid crystallizes in space group P1 with a = 4.706(1), b= 8.578(2), c= 16.521(3) ÅA; α= 85.9(1), β= 89.7(1)°, = 77.4(1). The crystal structure, solved by direct methods, has been refined to an R value of 0.046 for 2150 observed reflections. The two peptide molecules in the structure have somewhat extended conformations. The unlike molecules aggregate into separate alternating layers. Each layer is stabilized by hydrogen bonded head-to-tail sequences as well as sequences of hydrogen bonds involving peptide groups. The arrangement of molecules in each layer is similar to one of the plausible idealized arrangements of L-alanyl-L-alanine worked out from simple geometrical considerations. Adjacent layers in the structure are held together by interactions involving side chains as well as water molecules. The water structure observed in the complex provides a good model, at atomic resolution, for that in protein crystals. An interesting feature of the crystal structure is the existence of two water channels in the interfaces between adjacent peptide layers.

Interaction of Gibberellic Acid and Indole-3-acetic Acid in the Growth of Excised Cuscuta Shoot Tips in Vitro1

Gibberellic acid (GA3) induced a marked elongation of 2.5-centimeter shoot tips of Cuscuta chinensis Lamk. cultured in vitro. In terms of the absolute amount of elongation, this growth may be the largest reported for an isolated plant system. The response to hormone was dependent on an exogenous carbohydrate supply. The hormone-stimulated growth was due to both cell division and cell elongation. The growth response progressively decreased if GA3 was given at increasingly later times after culturing, but the decreased growth response could be restored by the application of indole-3-acetic acid (IAA) to the apex. Explants deprived of GA3 gradually lost their ability to transport IAA basipetally, but this ability was also restored by auxin application. The observations are explained on the basis that: (a) the growth of Cuscuta shoot tip in vitro requires, at least, both an auxin and a gibberellin; and (b) in the absence of gibberellin the cultured shoot tip explants lose the ability to produce and/or transport auxin.

Isonicotinic acid hydrazide - a reinvestigation

C6HvN30, orthorhombic, P2~2121, a = 14.915 (15), b=ll.400 (10), c=3.835 (5) A, Din= 1"417 (7), De= 1"395 g cm -3 and Z=4. The structure was refined by the least-squares method to an R of 0.072 for 699 observed reflexions. The angle between the mean planes of the pyridine ring and the acid hydrazide moiety is 18.1 °. The molecules are held together in the crystal by a network of N-H...N hydrogen bonds.

Role of glutamine synthetase in citric acid fermentation byAspergillus niger

The activity of glutamine synthetase fromAspergillus niger was significantly lowered under conditions of citric acid fermentation. The intracellular pH of the organism as determined by bromophenol blue dye distribution and fluorescein diacetate uptake methods was relatively constant between 6•0–6•5, when the pH of the external medium was varied between 2•3–7•0.Aspergillus niger glutamine synthetase was rapidly inactivated under acidic pH conditions and Mn2+ ions partially protected the enzyme against this inactivation. Mn2+-dependent glutamine synthetase activity was higher at acidic pH (6•0) compared to Mg2+-supported activity. While the concentration of Mg2+ required to optimally activate glutamine synthetase at pH 6•0 was very high (≥ 50 mM), Mn2+ was effective at 4 mM. Higher concentrations of Mn2+ were inhibitory. The inhibition of both Mn2+ and Mg2+-dependent reactions by citrate, 2-oxoglutarate and ATP were probably due to their ability to chelate divalent ions rather than as regulatory molecules. This suggestion was supported by the observation that a metal ion chelator, EDTA also produced similar effects. Of the end-products of the pathway, only histidine, carbamyl phosphate, AMP and ADP inhibitedAspergillus niger glutamine synthetase. The inhibitions were more pronounced when Mn2+ was the metal ion activator and greater inhibition was observed at lower pH values. These results permit us to postulate that glutamine synthesis may be markedly inhibited when the fungus is grown under conditions suitable for citric acid production and this block may result in delinking carbon and nitrogen metabolism leading to acidogenesis.

Phase transition in malonic acid: An infrared study

Malonic acid is shown to undergo an interesting phase transition at 360 K when the two non-equivalent cyclic hydrogen-bonded dimers present in the low-temperature phase become equivalent.

N-Carbamoyl-DL-aspartic acid

CsHaN205, PL a = 6.438 (2), b = 7.486 (3), c = 8.048 (4)A, a = 72.2(1), fl = 80.8(1), y = 76.4 (1) °, D m = 1.65 (1) (flotation), D c = 1.64 Mg m -3, Z = 2. Final R = 0.095 for 1205 observed reflections. The molecule assumes the sterically least favourable conformation with the side chain carboxyl group staggered between the a-carboxyl group and the N atom attached to C '~. The ureido group takes part in two specific interactions involving two nearly parallel hydrogen bonds in one and two convergent hydrogen bonds in the other.

Aspects of tautomerism 12-Some causes and consequences of participation in the solvolysis of acid chlorides

Solvolysis of nine representative half ester acid chlorides in aqueous acetone have been studied. Isomers solvolyse at distinctly different rates and furnish the original acids. Contrary to the well accepted views, no evidence for tautomerism or isomerism between the isomeric pairs of acid chlorides could be detected. In a number of cases alkoxy group participates in the solvolysis of neighbouring acid chlorides. This results in (a) rate enhancement and (b) partial or total shift of the reaction pattern from SN2 to SN1. Isomeric half ester acid chlorides, in the presence of a sufficiently strong Lewis acid, could give the same oxonium salt. Rearrangements observed in the reactions of unsymmetrical 1,2- and 1,3-dicarboxylic acid derivatives could be ascribed to the prior formation of common oxonium salt intermediates in the presence of Lewis acids.

Studies on the fractionation of total transfer ribonucleic acid and purification of an alanine transfer ribonucleic acid from chick embryo

Chick embryo tRNA, prepared by a simple large-scale method, was fractionated on three different ion-exchange columns. In all cases simple chromatographic patterns for various tRNA species were observed, indicating the presence of only a few major species of tRNA for each amino acid. By repeated chromatography one species of alanine tRNA was purified to approx. 80% purity. T1 ribonuclease digest of this purified tRNA gave a simple chromatographic pattern. Because of the simplicity of the method of preparation of tRNA from this readily available source and the presence of only a few species of tRNA for each amino acid, chick embryo is suited for the study of tRNA and its various functions in higher systems.

Infrared studies on the conformation of synthetic alamethicin fragments and model peptides containing .alpha.-aminoisobutyric acid

ABSTRACT: Infrared studies of synthetic alamethicin fragments and model peptides containing a-aminoisobutyric acid (Aib) have been carried out in solution. Tripeptides and larger fragments exhibit a strong tendency to form /3 turns, stabilized by 4 - 1 10-atom hydrogen bonds. Dipeptides show less well-defined structures, though C5 and C7 conformations are detectable. Conformational restrictions imposed by Aib residues result in these peptides populating a limited range of states. Integrated intensities of the hydrogen-bonded N-H stretching band can be used to quantitate the number of intramolecular hydrogen bonds. Predictions made from infrared data are in excellent agreement with nuclear magnetic resonance and X-ray diffraction studies. Assignments of the urethane and tertiary amide carbonyl groups in the free state have been made in model peptides. Shifts to lower frequency on hydrogen bonding are observed for the carbonyl groups. The 1-6 segment of alamethicin is shown to adopt a 310 helical structure stabilized by four intramolecular hydrogen bonds. The fragments Boc-Leu-Aib-Pro-Val-Aib-OMe (1 2-1 6) and Boc-Gly-Leu-Aib-Pro-Val-Aib-OMe (1 1-1 6) possess structures involving 4 - 1 and 5 - 1 hydrogen bonds. Supporting evidence for these structures is obtained from proton nuclear magnetic resonance studies.

Studies on crystalline complexes involving amino acids. V. The structure of L-serine-L-ascorbic acid

L-Serine-L-ascorbic acid, C3HTNOa. C6HsO6, a 1:1 complex between the amino acid serine and the vitamin ascorbic acid, crystallizes in the orthorhombic space group P2~2~2~ with four formula units in a cell of dimensions a = 5.335(3), b = 8.769(2), c = 25.782 (5) A. The structure was solved by direct methods and refined by full-matrix least squares to an R of 0.036 for 951 observed reflections. Both molecules are neutral in the structure. The conformation of the serine molecule is different from that observed in the crystal structures of L-serine, DL-serine and L-serine monohydrate. The enediol group in the ascorbic acid molecule is planar, whereas significant departures from planarity are observed in the lactone group. The conformation of this molecule is similar to that observed in arginine ascorbate. The unlike molecules aggregate into separate columns in the crystal structure. The columns are held together by hydrogen bonds. Among these, a pair of hydrogen bonds between the enediol group of ascorbic acid and the carboxylate group of serine provides a possible model for a specific interaction between ascorbic acid and a carboxylate ion.

Peptide models for the bacteriorhodopsin chromophore. Retinylidene-lysine systems containing aspartic acid and serine residues

The retinylidene Schiff base derivative of seven lysine containing peptides have been prepared in order to investigate solvent and neighboring group effects, on the absorption maximum of the protonated Schiff base chromophore. The peptides studied are Boc-Aib-Lys-Aib-OMe (1), Boc-Ala-Aib-Lys-OMe (2), Boc-Ala-Aib-Lys-Aib-OMe (3), Boc-Aib-Asp-Aib-Aib-Lys-Aib-OMe (4), Boc-Aib-Asp-Aib-Ala-Aib-Lys-Aib-OMe (5), Boc-Lys-Val-Gly-Phe-OMe (6) and Boc-Ser-Ala-Lys-Val-Gly-Phe-OMe (7). In all cases protonation shifts the absorption maxima to the red by 3150–8450 cm-1. For peptides 1–3 the protonation shifts are significantly larger in nonhydrogen bonding solvents like CHCl3 or CH2Cl2 as compared to hydrogen bonding solvents like CH3OH. The presence of a proximal Asp residue in 4 and 5 results in pronounced blue shift of the absorption maximum of the protonated Schiff base in CHCl3, relative to peptides lacking this residue. Peptides 6 and 7 represent small segments of the bacteriorhodopsin sequence in the vicinity of Lys-216. The presence of Ser reduces the magnitude of the protonation shift.

Unique presence of 2-methylthio-ribosylzeatin in the transfer ribonucleic acid of the bacterium Pseudomonas-Aeruginosa

Analysis of 35S labled nucleosides prepared from tRNA of Pseudomonas aeruginosa by phosphocellulose column chromatography, thin layer chromatography and Sephadex LH-20 column chromatography revealed the presence of 2-methylthioribosylzeatin in it. 2iPA, 6-(3-methyl-2-butenylamino)-9-β-D-ribofuranosyl purine; ms-2iPA, 6-(3-methyl-2-butenylamino)-2-methylthio-9-β-D-ribofuranosylpurine; ribosyl-cis-zeatin, 6-(4-hydroxy-3-methyl-cis-2-butenylamino)-9-β-D-ribofuranosylpurine; ribosyl-trans-zeatin, 6-(4-hydroxy-3-methyl-trans-2-butenylamino)-9-β-D-ribofuranosylpurine; ms-ribosylzeatin, 6-(4-hydroxy-3-methyl-2-butenylamino)-2-methylthio-9-β-D-ribofuranosylpurine; s4U2, 4-thiouridine; s2U*, 5-methylaminomethyl-2-thiouridine; s2C, 2-thiocytidine; TLC — thin layer chromatography.

Layer-by-layer self-assembly of modified hyaluronic acid/chitosan based on hydrogen bonding

The fabrication of hydrogen bonded polymer self-assembly for drug delivery has been accomplished via layer-by-layer sequential assembly from aqueous solution. In this study, the self-assembly was constructed based on hydrogen bonding between DNA base (adenine and thymine) pairs substituted on the backbone of chitosan and hyaluronic acid. Chitosan was modified with adenine, whereas hyaluronic acid was modified with thymine. Subsequently, these two polymers were sequentially absorbed on flat substrate by taking advantage of interactions of DNA base pairs via hydrogen bonding. Interlayer hydrogen bonding of these two polymers produces stable multilayer film without using any cross-linking agent. Thin film formation on quartz substrate has been monitored with UV-vis spectra and an AFM study. Formation of multilayer hydrogen-bonded thin film has been further confirmed with SEM. Encapsulation and release behavior of the therapeutic drug from the multilayer thin film at different conditions has been illustrated using UV-vis spectra. Cell viability of modified polymers using MTT assay confirmed no cytotoxic effect.

Importance of non-conserved distal carboxyl terminal amino acids in two peptidases belonging to the M1 family: Thermoplasma acidophilum Tricorn interacting factor F2 and Escherichia coli Peptidase N

Enzymes belonging to the M1 family play important cellular roles and the key amino acids (aa) in the catalytic domain are conserved. However, C-terminal domain aa are highly variable and demonstrate distinct differences in organization. To address a functional role for the C-terminal domain, progressive deletions were generated in Tricorn interacting factor F2 from Thermoplasma acidophilum (F2) and Peptidase N from Escherichia coli (PepN). Catalytic activity was partially reduced in PepN lacking 4 C-terminal residues (PepNΔC4) whereas it was greatly reduced in F2 lacking 10 C-terminal residues (F2ΔC10) or PepN lacking eleven C-terminal residues (PepNΔC11). Notably, expression of PepNΔC4, but not PepNΔC11, in E. coliΔpepN increased its ability to resist nutritional and high temperature stress, demonstrating physiological significance. Purified C-terminal deleted proteins demonstrated greater sensitivity to trypsin and bound stronger to 8-amino 1-napthalene sulphonic acid (ANS), revealing greater numbers of surface exposed hydrophobic aa. Also, F2 or PepN containing large aa deletions in the C-termini, but not smaller deletions, were present in high amounts in the insoluble fraction of cell extracts probably due to reduced protein solubility. Modeling studies, using the crystal structure of E. coli PepN, demonstrated increase in hydrophobic surface area and change in accessibility of several aa from buried to exposed upon deletion of C-terminal aa. Together, these studies revealed that non-conserved distal C-terminal aa repress the surface exposure of apolar aa, enhance protein solubility, and catalytic activity in two soluble and distinct members of the M1 family.