2' Derivatives of guanosine and inosine cyclic 3',5'-phosphates. Synthesis, enzymic activity, and the effect of 8-substituents.

A series of representative derivatives of guanosine cyclic 3',5'-phosphate (cGMP) and inosine cyclic 3',5'-phosphate (cIMP) which contained modifications in either the 2' position or the 8 and 2' positions were synthesized. Three types of derivatives were investigated: (1) derivatives in which the 2' position has been altered to produce a 2'-deoxynucleoside cyclic 3',5'-phosphate or a 9-beta-D-arabinofuranosylpurine cyclic 3',5'-phosphate; (2) 2'-omicron-acyl derivatives; and (3) doubly modified derivatives containing a 2' modification [as in (1) and (2)] and an 8-substitution. 2'-Deoxyinosine cyclic 3',5'-phosphate and 9-beta-D-arabinofuranosylhypoxanthine cyclic 3',5'-phosphate were obtained by HNO2 deamination of 2'-deoxyadenosine cyclic 3',5'-phosphate and 9-beta-D-arabinofuranosyladenine cyclic 3',5'-phosphate (ara-cAMP), respectively. Treatment of 8-bromo-2'-omicron-(p-toluenesulfonyl) adenosine cyclic 3',5'-phosphate with NaSH yielded the intermediate 8,2'-anhydro-9-beta-D-arabinofuranosyl-8-mercaptoadenine cyclic 3',5-phosphate, which was converted directly to 2'-deoxyadenosine cyclic 3',5'-phosphate (dcAMP) by treatment with Raney nickel. 8-Bromo-2'-omicron-(p-toluenesulfonyl) guanosine cyclic 3',5'-phosphate was converted to 8,2'-anhydro-9-beta-D-arabinofuranosyl-8-mercaptoguanine cyclic 3',5'-phosphate, and the latter was desulfurized with Raney nickel to give 2-deoxyguanosine cyclic 3',5'-phosphate. Ara-cAMP, 9-beta-D-arabinofuranosylguanine cyclic 3',5'-phosphate, and 9-beta-D-arabinofuranosyl-8-mercaptoguanine cyclic 3',5'-phosphate have been previously reported (Mian et al. (1974), J. Med. Chem. 17, 259). 8-Bromo-2'-omicron-acetylinosine cyclic 3',5'-phosphate and 8-[(p-chlorophenyl)thio]-2'-omicron-acetylinosine cyclic 3',5'-phosphate were produced by acylation of 8-bromoinosine cyclic 3',5'-phosphate and 8-[(p-chlorophenyl)thio]inosine cyclic 3',5'-phosphate, respectively; while 8-bromo-2'-omicron-butyrylguanosine cyclic 3',5'-phosphate was synthesized by bromination of 2'-omicron-butyrylguanosine cyclic 3',5'-phosphate.     

Phosphorus-31 nuclear magnetic resonance studies of the binding of nucleotides to NADP+-specific isocitrate dehydrogenase

The interaction of the 2'-phosphate-containing nucleotides (NADP+, NADPH, 2'-phosphoadenosine 5'-diphosphoribose, and adenosine 2',5'-bisphosphate) with NADP+ -specific isocitrate dehydrogenase was studied by using 31P NMR spectroscopy. The separate resonances corresponding to free and bound nucleotides, characteristic for slow exchange of nuclei on the NMR time scale, were observed in the spectra of the enzyme (obtained in the presence of excess ligand) with NADP+ and NADPH in the absence and presence of Mg2+ and with 2'-phosphoadenosine 5'-diphosphoribose in the absence of metal or in the presence of the substrate magnesium isocitrate. The position of the 31P resonance of the bound 2'-phosphate group in these spectra is invariant (delta = 6) in the pH range 5-8, indicating that the pK of this group is much lower in the complexes with the enzyme than that (pK = 6.13) in the free nucleotides. The additional downfield shift of this resonance by 1.8 ppm beyond that (delta = 4.22) of the dianionic form of the 2'-phosphate in free nucleotides suggests interaction with a positively charged group(s) and/or distortion of P-O-P angles as the result of binding to the enzyme. A single resonance of 2'-phosphate was observed in the spectrum of the enzyme complex with 2'-phosphoadenosine 5'-diphosphoribose in the presence of Mg2+, with the chemical shift dependent on the nucleotide to enzyme ratio, characteristic for the fast exchange situation. Addition of metal does not perturb the environment of the 2'-phosphate in the complexes of NADP+ and NADPH with isocitrate dehydrogenase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Conformationally locked GFP chromophore derivatives as potential fluorescent sensors

We propose to use the conformationally locked GFP chromophore derivatives (4-(2(difluoroboryl)benzylidene)-1H-imidazol-5(4H)-ones) as potential fluorescent sensors. The model carbamate derivatives were synthesized and their fluorescent properties were studied. It was shown that the introduction/cleavage of the carbamate groups at the nitrogen atom led to the noticeable shifts in the absorption and emission maxima of the resultant derivatives, which makes them promising candidates for new fluorescent sensors.     

Chemical synthesis of oligonucleotides containing a free sulphydryl group and subsequent attachment of thiol specific probes.

Oligonucleotides containing a free sulphydryl group at their 5'-termini have been synthesised and further derivatised with thiol specific probes. The nucleotide sequence required is prepared using standard solid phase phosphoramidite techniques and an extra round of synthesis is then performed using the S-triphenylmethyl O-methoxymorpholinophosphite derivatives of 2-mercaptoethanol, 3-mercaptopropan (1) ol or 6-mercaptohexan (1) ol. After cleavage from the resin and removal of the phosphate and base protecting groups, this yields an oligonucleotide containing an S-triphenylmethyl group attached to the 5'-phosphate group via a two, three or six carbon chain. The triphenylmethyl group can be readily removed with silver nitrate to give the free thiol. With the three and six carbon chain oligonucleotides, this thiol can be used, at pH 8, for the attachment of thiol specific probes as illustrated by the reaction with fluorescent conjugates of iodoacetates and maleiimides. However, oligonucleotides containing a thiol attached to the 5'-phosphate group via a two carbon chain are unstable at pH 8 decomposing to the free 5'-phosphate and so are unsuitable for further derivatisation.     

Spectrophotometric determination of pyridoxal and pyridoxal 5′-phosphate with 3-methyl-2-benzothiazolone hydrazone hydrochloride, and their selective assay

A spectrophotometric method with 3-methyl-2-benzothiazolone hydrazone hydrochloride was developed for the determination of pyridoxal and pyridoxal 5'-phosphate, and for the selective determination of each in the presence of the other. Pyridoxal and pyridoxal 5'-phosphate react with the reagent to yield the azine derivatives, which give characteristic absorption spectra. The highest extinction values are obtained when pyridoxal and pyridoxal 5'-phosphate are incubated at pH values of about 3.4 and 8.0 respectively; their maxima are at 430nm. (in 2.74x10(4)) and 380nm. (in 2.24x10(4)) respectively. The azine of pyridoxal is only slightly soluble under the neutral and alkaline conditions, whereas that of pyridoxal 5'-phosphate is substantially insoluble in the acid pH range. This difference in solubility of the azines made possible the selective determination of pyridoxal and pyridoxal 5'-phosphate. alpha-Oxoglutarate and pyruvate are among the substances shown not to interfere with the assay of pyridoxal; their derivatives absorb appreciably only at wavelengths below 420nm. For the assay of pyridoxal 5'-phosphate in the presence of these compounds measurement at 390nm. is necessary.     

Modification of mouse testicular lactate dehydrogenase by pyridoxal 5''-phosphate.

1. Mouse C4 lactate dehydrogenase treated in the dark with pyridoxal 5'-phosphate at pH8.7 and 25 degrees C loses activity gradually; 1mM-pyridoxal 5'-phosphate causes 83% inactivation, and higher concentrations of the reagent cause no further loss of activity. 2. The final extent of inactivation is very pH-dependent, greater inactivation occurring at the high pH values. 3. Inactivation may be fully reversed by addition of cysteine, or made permanent by reducing the enzyme with NaBH4. 4. The absorption spectrum of inactivated reduced enzyme indicates modification of lysine residues. Inactivation by 80% corresponds to modification of at least 1.8 mol of lysine/mol of enzyme subunit. 5. There is no loss of free thiol groups after inactivation with pyridoxal 5'-phosphate and reduction of the enzyme. 6. NAD+ or NADH gives complete protection against inactivation. protection studies with coenzyme fragments indicate that the AMP moiety is largely responsible for the protective effect. Lactate (10 mM) gives no protection in the absence of added nucleotides, but greatly enhances the protection given by ADP-ribose (1 mM). Thus ADP-ribose is able to trigger the binding of lactate. 7. Pyridoxal 5'-phosphate also acts as a non-covalent inhibitor of mouse C4 lactate dehydrogenase. The inhibition is non-competitive with respect to both NAD+ and lactate. 8. Km values for the enzyme at pH 8.0 and 25 degrees C, with the non-varied substrate saturating, are 0.3 mM-lactate and 5 microM-NAD+. 9. These results are discussed and compared with pyridoxal 5'-phosphate modification of other lactate dehydrogenase isoenzymes and related dehydrogenases.     

The nature of the circular-dichoric spectra of complexes between ribonuclease A and nucleotides.

The circular-dichroism and proton-magnetic-resonance spectra of complexes of ribonuclease A with dihydrouridine 3'-phosphate, 2'- and 3'-CMP, arabinosyl-3'-CMP, 1-(2-hydroxyethyl)cytosine 2'-phosphate and 1-(3-hydroxypropyl)cytosine 3'-phosphate were studied. Comparison of the results shows that non-additivity of the circular-dichroic spectrum of an enzyme-nucleotide complex may be due to: (a), alteration of the circular dichroic spectrum of the nucleotide under the influence of the asymmetric protein matrix (induced dichroism), and (b) a change in the nucleotide conformation. The contribution of each of the two factors was estimated to calculate the circular-dichoroic spectra of 2'-CMP and 3'-CMP in complex with ribonuclease A. 3'-CMP in this complex was characterized by negative circular dichroism in the long-wavelength absorption band of the nucleotide, whereas 2'-CMP was characterized by positive circular dichroism. Since both nucleotides in the complex are known to be in an anti conformation, it follows that even small changes in the conformation considerably modify the circular-dichroic spectrum of the nucleotide in complex with the enzyme.     

Effect of nucleotides on translocation of sugar nucleotides and adenosine 3'-phosphate 5'-phosphosulfate into Golgi apparatus vesicles

Recent studies from this laboratory have suggested that rat-liver Golgi apparatus derived membranes contain different proteins which can translocate in vitro CMP-N-acetylneuraminic acid, GDP-fucose and adenosine 3'-phosphate 5'-phosphosulfate from an external compartment into a lumenal one. The aim of this study was to define the role of the nucleotide, sugar and sulfate moieties of sugar nucleotides and adenosine 3'-phosphate 5'-phosphosulfate in translocation of these latter compounds across Golgi vesicle membranes. Indirect evidence was obtained suggesting that the nucleotide (but not sugar or sulfate) is a necessary recognition feature for binding to the Golgi membrane (measured as inhibition of translocation) but is not sufficient for overall translocation; this latter event also depends on the type of sugar. Important recognition features for inhibition of translocation of the above sugar nucleotides and adenosine 3'-phosphate 5'-phosphosulfate were found to be the type of nucleotide base (purine or pyrimidine) and the position of the phosphate group in the ribose. Thus, UMP and CMP were found to be competitive inhibitors of translocation of CMP-N-acetylneuraminic acid, while AMP did not inhibit. Structural features of the nucleotides which were less important in inhibition of translocation (and thus presumably in binding) of the above sugar nucleotides and adenosine 3'-phosphate 5'-phosphosulfate were the number of phosphate groups in the nucleotide (CDP and CMP inhibited to a similar extent), the presence of ribose or deoxyribose in the nucleotide, a replacement of hydrogen in positions 5 of pyrimidines or 8 in purines by halogens or an azido group. The sugar or sulfate did not inhibit translocation of the above sugar nucleotides and adenosine 3'-phosphate 5'-phosphosulfate into Golgi vesicles and therefore appear not to be involved in their binding to the Golgi membrane.     

Toxoplasma gondii: mechanism of the parasitostatic action of 6-thioxanthine

In contrast to the cytocidal effect of 6-thiopurines on mammalian cells, the action of 6-thioxanthine on Toxoplasma gondii was only parasitostatic. 6-Thioxanthine was a substrate of the parasite's hypoxanthine-guanine phosphoribosyltransferase. That enzyme converted 6-thioxanthine to 6-thioxanthosine 5'-phosphate which accumulated to near millimolar concentrations within parasites incubated intracellularly in medium containing the drug. 6-Thioxanthosine 5'-phosphate was the only detectable metabolite of 6-thioxanthine. The absence of 6-thioguanine nucleotides explains the lack of a parasitocidal effect because the incorporation of 6-thiodeoxyguanosine triphosphate into DNA is the mechanism of the lethal effect of 6-thiopurines on mammalian cells. Extracellular parasites that had accumulated a high concentration of 6-thioxanthosine 5'-phosphate incorporated more labeled hypoxanthine or xanthine into their nucleotide pools than did control parasites. The basis for this increased nucleobase salvage remains unexplained. It was not due to up-regulation of hypoxanthine-guanine phosphoribosyltransferase and could not be explained by reduced use of labeled nucleotides for nucleic acid synthesis. Extracellular parasites that had accumulated a high concentration of 6-thioxanthosine 5'-phosphate used labeled hypoxanthine almost entirely to make adenine nucleotides while control parasites made both adenine and guanine nucleotides. Both extracellular parasites that had accumulated a high concentration of 6-thioxanthosine 5'-phosphate and control parasites efficiently used labeled xanthine to make guanine nucleotides. These observations suggested that inosine 5'-phosphate-dehydrogenase was inhibited while guanosine 5'-phosphate synthase was not. Assay of inosine 5'-phosphate dehydrogenase in soluble extracts of T. gondii confirmed that 6-thioxanthosine 5'-phosphate was an inhibitor. We conclude that 6-thioxanthine blocks the growth of T. gondii by a depletion a guanine nucleotides.     

Chemical modification by pyridoxal 5''-phosphate and cyclohexane-1,2-dione indicates that Lys-7 and Arg-10 are involved in the p2 phosphate-binding subsite of bovine pancreatic ribonuclease A.

Steric and chemical evidence had previously shown that residues Lys-7 and/or Arg-10 of bovine pancreatic RNAase A could belong to the p2 phosphate-binding subsite, adjacent to the 3' side of the main site p1. In the present work chemical modification of the enzyme with pyridoxal 5'-phosphate and cyclohexane-1,2-dione was carried out in order to identify these residues positively as part of the p2 site. The reaction with pyridoxal 5'-phosphate yields three monosubstituted derivatives, at Lys-1, Lys-7 and Lys-41. A strong decrease in the yield of derivatives at Lys-7 and Lys-41 was observed when either p1 or p2 was specifically blocked by 5'-AMP or 3'-AMP respectively. These experiments indicate that both sites are needed for the reaction of pyridoxal 5'-phosphate with RNAase A to take place. The positive charge in one of the sites interacts with the phosphate group of pyridoxal 5'-phosphate, giving the proper orientation to the carbonyl group, which then reacts with the lysine residue present in the other site. The absence of reaction between pyridoxal 5'-phosphate and an RNAase derivative that has the p2 site blocked supports this hypothesis. Labelling of Lys-7 with pyridoxal 5'-phosphate has a more pronounced effect on the kinetics with RNA than with the smaller substrate 2',3'-cyclic CMP. In addition, when the phosphate moiety of the 5'-phosphopyridoxyl group was removed with alkaline phosphatase the kinetic constants with 2',3'-cyclic CMP returned to values very similar to those of the native enzyme, whereas a higher Km and lower Vmax. were still observed for RNA. This indicates that this new derivative has recovered a free p1 site and, hence, the capability to act on 2',3'-cyclic CMP, but the presence of the pyridoxyl group bound to Lys-7 is still blocking a secondary phosphate-binding site, namely p2. Finally, reaction of cyclohexane-1,2-dione at Arg-10 is suppressed in the presence of 3'-AMP but only a 19% decrease is observed with 5'-AMP, suggesting that Arg-10 is also close to the p2 phosphate-binding subsite.     

Chemical synthesis of 5''-pyrophosphate and triphosphate derivatives of 3''-5'' ApA, ApG, GpA and GpG. CD study of the effect of 5''-phosphate groups on the conformation of 3''-5'' GpG.

A simple, two-step method is described for the synthesis of the 5'-pyro- and triphosphate derivatives of 3'-5' ApA, ApG, GpA and GpG. The readily accessible 2'(3')-5' ApA, ApG, GpA and GpG were converted in one step to the corresponding 5'-phosphoramidate derivatives which were then transformed to the 5'-pyro- and triphosphates. CD spectra of 3'-5' pn GpG (n = 0,1,2 or 3) derivatives, measured at pH 1, indicated stabilization of the (syn) G+p (anti)G conformation by the 5'-phosphate groups.     

Phospholipase C from Bacillus cereus. Evidence for essential lysine residues.

1. Phospholipase C was inactivated by exposure to the three amino-group reagents, ethyl acetamidate, 2,4,6-trinitrobenzensulphonic acid and pyridoxal 5'-phosphate plus reduction. 2. Inactivation by pyridoxal 5'-phosphate showed the characteristics of Schiff's base formation with the enzyme. The pyridoxal 5'-phosphate-treated enzyme after reduction had an absorbance maximum at 325 mm and 6-N-pyridoxyl-lysine was the only fluorescent component after acid hydrolysis. 3. For complete inactivation, 2 mol of pyridoxal 5'-phosphate or 7 mol of 2,4,6-trinitrophenyl were incorporated/mol of enzyme. 4. The two apparently essential lysine residues were much more reactive to pyridoxal 5'-phosphate than the other 19 lysine residues in the enzyme. 5. Binding of phospholipase C to a substrate-based affinity gel caused marked protection against inactivation by pyridoxal 5'-phosphate. For complete inactivation of the gel-bound enzyme, 5 mol of pyridoxal 5'-phosphate were incorporated/mol of enzyme and there was no evidence of two especially reactive lysine residues. 6. On application of pyridoxal 5'-phosphate-treated enzyme (remaining activity 30% of original) to a column of the affinity gel, some material bound and some did not. The latter contained very little enzyme activity and was heavily incorporated with reagent (9.06 mol/mol of enzyme). The former had a specific activity of 34% of that of the control and contained 1.29 mol of reagent/mol of enzyme. 7. Thus phospholipase C appears to contain two lysine residues that are essential for enzyme activity, but probably not for substrate binding.     

Fluorescent derivatives of bile salts. I. Synthesis and properties of NBD-amino derivatives of bile salts.

In order to visualize bile salt transport, fluorescent bile salt derivatives were synthesized by introduction of the relatively small fluorescent 4-nitrobenzo-2-oxa-1,3-diazol (NBD)-amino group in either the 3-, 7-, or 12-position of the steroid structure, thus providing a complete set of diastereomeric derivatives, 3 alpha-NBD-amino-7 alpha,12 alpha-dihydroxy-5 beta-cholan-24-oic acid, 3 beta-NBD-amino-7 alpha,12 alpha-dihydroxy-5 beta-cholan-24-oic acid, 7 alpha-NBD-amino-3 alpha,12 alpha-dihydroxy-5 beta-cholan-24-oic acid, 7 beta-NBD-amino-3 alpha,12 alpha-dihydroxy-5 beta-cholan-24-oic acid, 12 alpha-NBD-amino-3 alpha,7 alpha-dihydroxy-5 beta-cholan-24-oic acid, 12 beta-NBD-amino-3 alpha,7 alpha-dihydroxy-5 beta-cholan-24-oic acid, as well as their taurine conjugates. Their optical properties with absorption maxima at about 490 nm and emission maxima at 550 nm make them suitable for fluorescent microscopic studies. Fluorescence of the NBD-derivatives is strongly dependent on polarity of the solvent, on the concentration of the bile salt derivatives, and only slightly on temperature.     

The Sonogashira reaction as a new method for the modification of borated analogues of the green fluorescence protein chromophore

The Sonogashira reaction was used for modifications of borated green fluorescence protein chromophore derivatives, 4-(2-(difluoroboryl)benzylidene)-1H-imidazol-5(4H)-ones, for the development of new fluorescent dyes. The derivatives bearing an acetylene fragment and a difluoroboryl group were obtained in high yields. The modification resulted in a significant bathochromic shift of the absorption and emission maxima and is a promising method for the development of new fluorescent dyes.     

Custom fluorescent-nucleotide synthesis as an alternative method for nucleic acid labeling

The variety of potentially useful dyes or haptenes available for fluorescent nucleic acid hybridization assays is far greater than what can be obtained from commercial sources. Since this diversity could be useful in many laboratory applications, we have developed a simple and inexpensive procedure for preparing nonpurified labeled nucleotides, for use in common nucleic acid labeling reactions, such as PCR and nick translation. The modified nucleotides were synthesized by coupling allylamine-dUTP to the succinimidyl-ester derivatives of the fluorescent dyes or haptenes such as biotin or digoxigenin, which require fluorescently labeled proteins for detection. This method allows custom preparation of most common fluorescent nucleotides and rapid testing of new ones, while reducing the cost of procedures such as multiplex fluorescent in situ hybridization (M-FISH) by 100-200 fold.     

Microbiology of Saturated Salt Solutions and Other Harsh Environments. IV. New Observations of Ribonucleotide-Induced Recovery of KCl-Habituated Penicillium notatum

Salt-tolerant mutant Penicillium notatum sub-cultured in a glucose-peptone broth saturated with KCl shows continued attenuated growth when transferred to salt-free broth. Additional tests have shown E. coli S-RNA to be inferior to yeast RNA preparations, that base-free phosphate sources are inactive, but that nicotinamide adenine dinucleotide and flavine adenine dinucleotide are moderately active. All phosphate derivatives of adenine, cytosine and guanosine and inosine were active including 5'-polyphosphates, 3'(2')-monophosphates 5'-monophosphates, and adenine 3', 5'-cyclic monophosphate. Uracil derivatives were of low activity at best.Among base precursors, orotic acid was moderately active whereas imidazoles were not. The high activity of inosine 5'-phosphate a precursor of other purine nucleotides suggested that one mode of KCl action might involve a block in conversion of 4-amino-5-imidazole carboxamide ribonucleoside to the hypoxanthine nucleotide.     

Footprinting of Chlorella virus DNA ligase bound at a nick in duplex DNA.

The 298-amino acid ATP-dependent DNA ligase of Chlorella virus PBCV-1 is the smallest eukaryotic DNA ligase known. The enzyme has intrinsic specificity for binding to nicked duplex DNA. To delineate the ligase-DNA interface, we have footprinted the enzyme binding site on DNA and the DNA binding site on ligase. The size of the exonuclease III footprint of ligase bound a single nick in duplex DNA is 19-21 nucleotides. The footprint is asymmetric, extending 8-9 nucleotides on the 3'-OH side of the nick and 11-12 nucleotides on the 5'-phosphate side. The 5'-phosphate moiety is essential for the binding of Chlorella virus ligase to nicked DNA. Here we show that the 3'-OH moiety is not required for nick recognition. The Chlorella virus ligase binds to a nicked ligand containing 2',3'-dideoxy and 5'-phosphate termini, but cannot catalyze adenylation of the 5'-end. Hence, the 3'-OH is important for step 2 chemistry even though it is not itself chemically transformed during DNA-adenylate formation. A 2'-OH cannot substitute for the essential 3'-OH in adenylation at a nick or even in strand closure at a preadenylated nick. The protein side of the ligase-DNA interface was probed by limited proteolysis of ligase with trypsin and chymotrypsin in the presence and absence of nicked DNA. Protease accessible sites are clustered within a short segment from amino acids 210-225 located distal to conserved motif V. The ligase is protected from proteolysis by nicked DNA. Protease cleavage of the native enzyme prior to DNA addition results in loss of DNA binding. These results suggest a bipartite domain structure in which the interdomain segment either comprises part of the DNA binding site or undergoes a conformational change upon DNA binding. The domain structure of Chlorella virus ligase inferred from the solution experiments is consistent with the structure of T7 DNA ligase determined by x-ray crystallography.     

Conformational analysis of canonical 2-deoxyribonucleotides. 2. Purine nucleotides

The molecular structure of different conformers of isolated canonical purine 2'-deoxyribonucleotides 2-deoxyadenosine-5'-phosphate (pdA) and 2'-deoxyguanosine-5'-phosphate (pdG) was optimized using the B3LYP/6-31G(d) method. The results of the calculations reveal that the geometrical parameters and relative stability of the conformers significantly depend on the nature of the nucleobase, its orientation, the conformation of the furanose ring, the charge of the phosphate group and the character of the intramolecular hydrogen bonds. Analysis of the electron density distribution in purine nucleotides reveals the existence of a number of intramolecular hydrogen bonds. In general, the south conformer has a lower energy at the anti orientation of the base, and both conformers occur as the most stable for the syn orientation of the nucleobases. The results of the calculations reveal that the geometry and relative energy of the conformers of purine DNTs may be easily tuned by the charge of the phosphate group.     

The chemotactic response of the myxomycete Physarum polyhcepalum to amino acids, cyclic nucleotides and folic acid

Abstract The plasmodium of Physarum polycephalum exhibited positive chemotaxis towards l -alanine, l -aspartate, l -asparagine, l -glutamate, glycine, l -leucine, l -serine, and l -threonine and negative chemotaxis towards l -tryptophan. All attractant amino acids, except l -serine and l -threonine competed with each other; l -serine and l -threonine competed with the other amino acids but did not interfere with the response to each other. Cyclic nucleotides were attractants and cyclic 3',5'- or 2',3'-phosphate derivatives of either adenine or guanine were active, wheras compounds lacking the ring structure generally were not. Folic acid was an attractant whereas certain aromatic compounds were either inactive or repellent.