Radiation damage in solid 5-halouracils: free radicals in single crystals of 5-fluorouracil

X-irradiation at 300 K of single crystals of 5-fluorouracil results in the formation of two different radical species observable by e.s.r. and ENDOR-spectroscopy. One is an alpha-fluoro radical RCF(CH2)R' formed by saturation of the 5,6-double bond of the pyrimidine ring. The principal values of its alpha-fluorine interaction are 170, -9 and -18 G; the isotropic part of the methylene beta-proton couplings are 46.3 and 28 G, respectively; the g-tensor has principal values of 2.0029, 2.0066, and 2.0052. The other radical species is formed by enolization of the C4-carbonyl function. The resulting spin-density distribution gives rise to three observable interactions, an alpha-proton (-5.2, -14.9, -11.2 G) at C6, an OH-proton from the C4--OH group (-1.7, -5.0, -4.1 G) and a residual alpha-fluorine interaction (0,0,11.2G). Irradiation at 77 K yields an e.s.r.-pattern which is tentatively assigned to the molecular anion radical. These findings are related to the radiation chemistry of solid 5-halouracils.     

Phenylalanyl-tRNA synthetase of Escherichia coli K 10. Multiple enzyme-aminoacyl-tRNA complexes as a consequence of substrate specificity.

The interaction between Phe-tRNA(Phe) or other acyl-tRNA derivatives thereof and phenylalanyl-tRNA synthetase of Escherichia coli K 10 has been investigated by nonequilibrium dialysis, by fluorescence titration in the presence of 2-p-toluidinylnaphthalene-6-sulfonate, by the kinetics of the aminoacylation of tRNA(Phe), and by the kinetics of the catalytic hydrolysis of Phe-tRNA(Phe). Phe-tRNA(Phe), or derivatives thereof, forms two types of complexes with the synthetase. One type involves the attachment of the phenylalanyl moiety to the phenylalanine-specific site of the enzyme, and the other type, to the tRNA(Phe)-specific binding site. They resemble alternative modes of a destabilized enzyme-product complex and are predicted on the basis of thermodynamic considerations. The two modes of binding of acyl-tRNA compete with each other. The attachment of Phe-tRNA(Phe) to the phenylalanine-specific site dominates. At equilibrium, this complex is present at a fourfold higher concentration than the other type of complex. The HNO2 deaminated Phe-tRNA(Phe) binds exclusively to the site specific for L-phenylalanine. On the contrary, Ile-tRNA(Phe) adds at 94.1% to the tRNA(Phe)-specific site. The association of Phe-tRNA(Phe) with this site leads to enzymatic hydrolysis into L-phenylalanine and tRNA(Phe). The complex involving the phenylalanine-specific site is hydrolytically unproductive. L-Phenylalanine acts as an activator of the hydrolysis by occupying the amino acid specific site and by shifting the equilibrium between the complexes toward the binding ot Phe-tRNA(Phe) at the tRNA(Phe)-specific site. The association of Phe-tRNA(Phe) at the phenylalanine-specific site does not interfere sterically with the binding of free tRNA(Phe). The sequential addition of free and aminoacylated tRNA(Phe) exhibits negative cooperativity. Such a mechanism could help to expel the product from the enzyme.     

Variable pH dependence of the myosin-ATPase in different muscles of the rat

Summary For the histochemical demonstration of the Myosin-ATPase (EC 3.6.1.3) the pH of both the preincubation and the incubation medium was varied in steps of 1 within a small range: 10.2 to 10.5 and 9.3 to 9.6, respectively. The optimum combinations of both pH values, defined as the ones providing most consistent contrast among the three major types of muscle fibers were determined in 9 different muscles of the rat. The spectrum of optimum combinations differs considerably from muscle to muscle. The reduction of the incubation pH by only 0.1 may drastically change the staining pattern. This probably reflects the unspecifity of the histochemical procedure as well as the plasticity of the ATPase systems. To cope with the lability of the myosin-ATPase the optimum pH values of both media should be determined for each muscle separately.     

Irradiation of micro-organisms with mono-energetic X-rays; biological consequences of the Auger effect.

The radiation resonance effect reported previously for isolated biomolecules has now for the first time been observed in a cellular system. Dried bacteria, Micrococcus denitrificans, in which TdR in DNA was partially substituted by BUdR, were subjected to mono-energetic X-rays of energies below or above the K-edge for Br. Subsequently, the colony-forming ability was assayed. For photon energy slightly above the K-edge, the lethality/rad was greater than that below the K-edge. This is interpreted in terms of the Auger effect initiated selectively by photo-absorption in constituent Br atoms. The differential absorption of low-energy photons in constituent atoms of DNA is also discussed.     

Influence of the hormone analogue 13-NLE-molitin and of 1-methyl-3-isobutylxanthine on tone and cyclic 3',5'-AMP content of antral and duodenal muscles in the rabbit.

1. By the action of 1-methyl-3-isobutylxanthine (isobutyltheophylline, 2 - 3 × 10⁻⁴ M), the content of cyclic 3', 5'-AMP in the antral and duodenal muscles of the rabbit is increased by 72 % and 126 %, respectively; by 1.8 × 10⁻⁷ M 13-norleucine-motilin and 1.8 × 10⁻⁶ M acetylcholine it is not changed. 13-norleucine-motilin is an analogue of the recently discovered duodenal tissue hormone motilin and has identical effects. 1-methyl-3-isobutylxanthine has a more powerful inhibiting effect on phosphodiesterase than has theophylline.2. 3 × 10⁻⁴ M isobutyltheophylline reduces the tone of the duodenal muscle while simultaneously increasing the content of cyclic AMP and negates the tone-enhancing effect of nle-motilin on the duodenal muscle, while nle-motilin increases the muscle tone lowered by isobutyltheophylline.3. The basic tone of the antral muscle is not reduced by isobutyltheophylline. However, the contraction-promoting effect of nle-motilin after an increase in cyclic AMP due to isobutyltheophylline is significantly lower.4. It is assumed that the changes in the tone or in the response of the antral and duodenal muscles to nle-motilin observed after the administration of isobutyltheophylline, are due to the increase of cyclic AMP in the tissue.5. The antagonistic effects of cyclic AMP and motilin on the gastro-intestinal muscles might be of physiological importance for the regulation of the gastro-intestinal motor activity.     

Isolation and characterization of cDNA clones encoding a 18.8 kDa polypeptide,the product of the gene psaL,associated with photosystem I reaction center from spinach

Several cDNA clones encoding subunit XI of photosystem I reaction center (PSI-L) have been isolated from two gt11 expression libraries based on polyadenylated RNA of spinach seedlings illuminated for 4 and 16 h, respectively. The precursor polypeptide made from these recombinant DNAs in vitro can be efficiently imported into isolated spinach chloroplasts. It is correctly processed to the size of the authentic polypeptide and integrates into the photosystem I assembly. The 834 nucleotide sequence of the longest cDNA insert encodes a precursor polypeptide of 24 kDa (216 residues) and a mature protein of probably 18.8 kDa (169 residues). Hydropathy analysis suggests that the polypeptide contains two transmembrane segments. The protein appears to originate in a single-copy gene in spinach and to be decoded from RNA species of ca. 900 bases.