Molecular mechanism for the specific inhibition of reverse transcriptase of rous sarcoma virus by the copper complexes of isonicotinic acid hydrazide

Isonicotinic acid hydrazide (isoniazid), one of the most potent antitubercular drugs, was recently shown, in our laboratory, to form two different complexes with copper, depending upon the oxidation state of the metal ion. Both the complexes have been shown to possess antiviral activity against Rous sarcoma virus, an RNA tumor virus. The antiviral activity of the complexes has been attributed to their ability to inhibit the endogenous reverse transcriptase activity of RSV. More recent studies in our laboratory indicate that both these complexes inhibit both endogenous and exogenous reactions. As low a final concentration as 50 μM of the cupric and the cuprous complexes inhibits the endogenous reaction to the extent of 93 and 75 per cent respectively. Inhibition of the exogenous reaction varies with the templates. The inhibition can be reversed by either β-mercaptoethanol or ethylene-diamine-tetra-acetic acid. The specificity of this inhibition has been ascertained by using a synthetic primer-template, −(dG)not, vert, similar15−(rCm)n, which is highly specific for reverse transcriptases. The inhibition is found to be template specific. The studies carried out, using various synthetic primer-templates, show the inhibition of both the steps of reverse transcription by the copper complexes of isoniazid.

Effect of depletion of vitamin A, followed by supplementation with retinyl acetate or retinoic acid, on regeneration of rat liver

After partial hepatectomy the net increase in tissue weight and in RNA, DNA and proteins in the regenerating liver was markedly less in vitamin A-deleted or retinoic acid-supplemented male rats, compared with the corresponding normal control or retinyl acetate-supplemented ones.

A Case Study of the Conformation of Poly(alpha-aminoisobutyric acid): alpha- or 310-Helix

The relative stabilities of a- and Blo-helical structures for polymers of a-aminoisobutyric acid (Aib) have been worked out, using the classical potential energy functions. To make a comparative study, we have used Buckingham "6-exp" and Kitaigorodsky's potential functions. Conformational analysis of the dipeptide segment with Aib residue indicates the necessity for nonplanar distortion of the peptide unit, which is a common feature in the observed crystal structures with Aib residues. In the range of Aw -10 to +loo studied, a-helical conformations are preferred in the region -3" < Aw < +loo, and Blo-helical conformations are preferred in the region -3" > Aw > -10'. Minimum energy conformations for right-handed structures are found in the +ue region of Aw and correspondingly for left-handed structures in the -ue region of Aw. For Aw - 6", a-helical structures have four- or near fourfold symmetry with h - 1.5 A. Such a helix with n = 4 and h = 1.5 A is termed an a'-helix. This structure is found to be consistent with the electron diffraction data of Malcolm3 and energetically more favorable than the standard 310-helix.

Rate of ribonucleic acid chain growth in Mycobacterium tuberculosis H37Rv.

Two methods were employed to measure the rate of ribonucleic acid (RNA) chain growth in vivo in Mycobacterium tuberculosis H37Rv cultures growing in Sauton medium at 37 degrees C, with a generation time of 10 h. In the first, the bacteria were allowed to assimilate [3H]uracil or [3H]guanine into their RNA for short time periods. The RNA was then extracted and hydrolyzed with alkali, and the radioactivity in the resulting nucleotides and nucleosides was measured. The data obtained by this method allowed the calculation of the individual nucleotide step times during the growth of RNA chains, from which the average rate of RNA chain elongation was estimated to be about 4 nucleotides per s. The second method employed the antibiotic rifampin, which specifically inhibits the initiation of RNA synthesis without interfering with the elongation and completion of nascent RNA chains. Usint this method, the transcription time of the 16S, 23S, and 5S ribosomal RNA genes was estimated to be 7.6 min, which corresponds to a ribosomal RNA chain growth rate of 10 nucleotides per s.

p-hydroxymandelic acid - a key intermediate in the metabolism of DL(+)-phenylalanine by aspergillus niger

The metabolism of phenylalanine by a strain of Aspergillus niger, isolated from the soil by enrichment culture has been studied. Analyses of the culture filtrates and replacement studies with various metabolites have revealed the operation of a degradative pathway involving p-hydroxymandelate as a key intermediate in this organism, p-Hydroxymandelate has been isolated from the cultural filtrates and its identity established by UV, IR and chromatographic techniques. A scheme for the degradation of phenylalanine in this organism has been proposed.

At4g24160, a Soluble Acyl-Coenzyme A-Dependent Lysophosphatidic Acid Acyltransferase

Human CGI-58 (for comparative gene identification-58) and YLR099c, encoding Ict1p in Saccharomyces cerevisiae, have recently been identified as acyl-CoA-dependent lysophosphatidic acid acyltransferases. Sequence database searches for CGI-58 like proteins in Arabidopsis (Arabidopsis thaliana) revealed 24 proteins with At4g24160, a member of the alpha/beta-hydrolase family of proteins being the closest homolog. At4g24160 contains three motifs that are conserved across the plant species: a GXSXG lipase motif, a HX4D acyltransferase motif, and V(X)(3)HGF, a probable lipid binding motif. Dendrogram analysis of yeast ICT1, CGI-58, and At4g24160 placed these three polypeptides in the same group. Here, we describe and characterize At4g24160 as, to our knowledge, the first soluble lysophosphatidic acid acyltransferase in plants. A lipidomics approach revealed that At4g24160 has additional triacylglycerol lipase and phosphatidylcholine hydrolyzing enzymatic activities. These data establish At4g24160, a protein with a previously unknown function, as an enzyme that might play a pivotal role in maintaining the lipid homeostasis in plants by regulating both phospholipid and neutral lipid levels.

Conformational Choices for the Stereochemically Constrained gamma-Amino Acid Residue Gabapentin: Theoretical Studies and Correlation With Experimental Results

Gabapentin (1-aminomethylcyclohexaneacetic acid, Gpn) is an achiral, conformationally constrained gamma amino acid residue. A survey of available crystal structures of Gpn peptides reveals that the torsion angles about the C-gamma-C-beta (theta(1)) and C-beta-C-alpha(theta(2)) bonds are overwhelmingly limited to gauche, gauche (g(+)g(+)/g(-)g(-)) conformations. The Gpn residue forms C-7 and C-9 hydrogen bonds in which the donor and acceptor atoms come from the flanking peptide units. In combination with alpha amino acid residues alpha gamma and gamma alpha segments can adopt C-12 hydrogen bonded structures. The conformational choices available to the Gpn residue have been probed using energy calculations, adopting a grid search strategy. Ramachandran phi-psi maps have been constructed for fixed values of theta(1) and theta(2), corresponding to the gauche and trans conformations. The sterically allowed and energetically favorable regions of conformational space have been defined and experimental observations compared. C-7 and C-9 hydrogen bonded conformational families have been identified using a grid search approach in which theta(1) and theta(2) values are varied over a range of +/- 10 degrees about ideal values at 1 degrees intervals. The theoretical analysis together with experimental observations for 59 Gpn residues from 35 crystal structures permits definition of the limited range of conformational possibilities at this gamma amino acid residue. .

Stereochemical restrictions on the occurrence of amino acid residues in the collagen structure

The primary structure of collagen is characterized by the repeating tripeptide sequence (Gly-R2-R3)n. The results of theoretical studies, carried out using contact criteria to compute the stereochemically allowed orientations for various side chains at locations 2 and 3, are reported here. It is found that side chains with only γ-atoms, as in valine, serine and threonine, or with only one δ-methyl group, as in isoleucine, can occur equally well at locations 2 and 3, as is actually the case in collagen. Side chains with two Cδ-atoms, as in leucine and phenyl-alanine, can also be accommodated at both positions. However, if they occur as R3 their freedom of orientation is severely restricted in the presence of a proline residue as R2 in a neighbouring chain. If water molecules bound to the chains of the triple helix are assumed to be present, then location 3 is virtually impossible for leucine and phenylalanine residues. Location 2 is, however, unaffected, and their presence as R2 can help to shield the water molecules from disturbance by the solvent medium. This may be the reason for the preferential occurrence of Leu and Phe residues in location 2 in the collagen triplets, although the polypeptides (Gly-Pro-Leu)n and (Gly-Pro-Phe)n form collagen-like structures.

Mechanism of action of selenious acid in the electrodeposition of manganese

The effect of selenious acid as an addition agent in the electrodeposition of manganese was studied by analysing the current-potential curves for manganese deposition. The mechanism of action of this addition agent was found to be essentially similar to that proposed for sulphur dioxide, namely to affect the manganese deposition indirectly by influencing the hydrogen evolution reaction which is a parallel reaction at the electrode surface.

A novel amino acid racemization in vitamin b6-amino acid schiff base copper complexes: crystal structures of aquo (5'-phosphopyridoxylidene-dl-tyrosinato) copper(II) 3.5H2O and aquo (5'-phosphopyridoxylidene-dl-phenylalaninato) copper(II) 2.5H2O

A novel racemization observed in the Vitamin B6-amino acid Schiff base complexes, aquo (5'-phosphopyridoxylidene-l-tyrosinato) copper(II) and aquo (5'-phosphopyridoxylidene-l-phenylalaninato) copper(II) is described. The racemization taking place in solution under mild acidic conditions (pH 5-6) was confirmed by CD studies and the products were characterized by single crystal X-ray diffraction. The structures of both complexes show almost parallel orientation of the aromatic side chain and the pyridoxal II-system. The activation of the αCsingle bondH group due to the intermolecular II- interaction is probably the reason for the unusual racemization observed.

X-ray studies on crystalline complexes involving amino acids and peptides. XXIII. Variability in ionization state, conformation and molecular aggregation in the complexes of succinic acid with DL- and L-lysine

Crystalline complexes of succinic acid with DL- and L-lysine have been prepared and analysed by X-ray diffraction. DL-Lysine complex: C6HIsN202 + 1 2- 1 ~C4H404 .~C4H604, Mr -- 264"2, PI, a = 5"506 (4), =8.070(2), c=14.089(2) A,, a=92.02(1), /3= 100"69 (3), y = 95"85 (3) ~>, Z = 2, Dx = 1"44 g cm -3, R = 0.059 for 2546 observed reflections. Form I of the e-lysine complex: C6HIsN20-, ~ .C4H504, Mr = 264.2, P1, a = 5" 125 (2), b = 8"087 (1), c = 8"689 (1) A,, a = 112.06 (1), /3 = 99.08 (2), y = 93"77(2) °, Z--l, D,,,=1"34(3), Dx=l"34gcm 3 R = 0.033 for 1475 observed reflections. Form II of + I 2- the e-lysine complex: C6H15N202 .,iC4H404 .- 1 I ") 4C4H604.4(C4HsO4""H'"CaH404)" , Mr = 264"2, P1, a = 10.143 (4), b = 10.256 (2), c = 12"916 (3) A,, a = 105.00 (2),/3 = 99-09 (3), y = 92"78 (3)::, Z = 4, Dm= 1"37(4), D,.= 1.38gcm 3, R=0.067 for 2809 observed reflections. The succinic acid molecules in the structures exhibit a variety of ionization states. Two of the lysine conformations found in the complexes have been observed for the first time in crystals containing lysine. Form II of the L-lysine complex is highly pseudosymmetric. In all the complexes, unlike molecules aggregate into separate alternating layers. The basic element of aggregation in the lysine layer in the complexes is an S2-type head-to-tail sequence. This element combines in different ways in the three structures. The basic element of aggre gation in the succinic acid layer in the complexes is a hydrogen-bonded ribbon. The ribbons are interconnected indirectly through amino groups in the lysine layer.

A comparative study of the early effects of phenobarbital and 3-methylcholanthrene on the synthesis and transport of ribonucleic acid in rat liver.

At 2-3 h after phenobaribtal administration, the drug has no effect on nucleoplasmic RNA synthesis and decreases nucleolar RNA synthesis. However, at this time there is an increase in the labelling of cytoplasmic poly(A)-containing RNA, even though there is decreased labelling of total polyribosomal RNA. The decrease in labelling of nucleolar and total polyribosomal RNA owing to phenobarbital is a transient phenomenon. Under similar conditions, 3-methylcholanthrene has no effect on nucleolar RNA synthesis, but leads to an increase in synthesis of nucleoplasmic and cytoplasmic poly(A)-containing RNA. Cytosol isolated from phenobarbital-treated, but not from 3-methyl-cholanthrene-treated, animals facilitates an enhanced transport of RNA from nuclei. At the time points investigated, 3-methylcholanthrene or its metabolite shows a 10-15-fold higher concentration in the chromatin than that of phenobarbital or its metabolite. It is suggested that the primary effect of phenobarbital is at the cytoplasmic level, promoting the transport of RNA from the nuclei, which can act as a trigger for enhanced transcription at later periods. 3-Methylcholanthrene or its metabolite directly binds to the chromatin and evokes a selective transcriptional response.

Effect of Nucleic Acid Reactive Antibodies on Tumor Cells Grown in Vivo

Nucleic acid reactive antibodies have been reported to inhibit various nucleio acid mediated functions in cell free systems. These antibodies were also shown to inhibit the growth of transformed cells in culture due to the high rate of endocytosis in transformed cells as compared to normal cells. In this report, we have tested the possibility of nucleic acid reactive antibodies inhibiting the growth of tumor cells in vivo. The life span of mice bearing Dalton's lymphoma ascites tumor cells was increased, when they were immunized with conjugates of guanosine-BSA, GMP-BSA and tRNA-MBSA complex before transplanting the tumor cells. A similar effect was also observed when mice were injected intraperitoneally with antibodies to guanosine oi GMP along with the tumor cells. The specificity was ascertained, as immunization with non-specific antigens did not show any significant effect on tumor bearing mice. The results shows that nucleic acid. reactive antibodies inhibit the growth of tumor cells in vivo.

Aspects of tautomerism. 8. Solvolysis of some pseudo and normal acid chlorides

Pseudo acid chlorides derived from levulinic acid ando-benzoyl-benzoic acid, solvolyse in aqueous acetone, aqueous dioxane and aqueous dimethylformamide by aS Nl process. Their reaction pattern is distinct from that of typical normal acid chlorides, viz.,p-benzoylbenzoyl chloride and fluorene-9-one-1-carboxylic acid chloride, which solvolyse by aS N2 pathway. No evidence for tautomerism could be obtained either between the normal and pseudo forms of the acid chlorides or the derived ion pairs.