DNA G-quartets in a 1.86 A resolution structure of an Oxytricha nova telomeric protein-DNA complex.

The Oxytricha nova telomere end binding protein (OnTEBP) recognizes, binds and protects the single-stranded 3'-terminal DNA extension found at the ends of macronuclear chromosomes. The structure of this complex shows that the single strand GGGGTTTTGGGG DNA binds in a deep cleft between the two protein subunits of OnTEBP, adopting a non-helical and irregular conformation. In extending the resolution limit of this structure to 1.86 A, we were surprised to find a G-quartet linked dimer of the GGGGTTTTGGGG DNA also packing within the crystal lattice and interacting with the telomere end binding protein. The G-quartet DNA exhibits the same structure and topology as previously observed in solution by NMR with diagonally crossing d(TTTT) loops at either end of the four-stranded helix. Additionally, the crystal structure reveals clearly visible Na(+), and specific patterns of bound water molecules in the four non-equivalent grooves. Although the G-quartet:protein contact surfaces are modest and might simply represent crystal packing interactions, it is interesting to speculate that the two types of telomeric DNA-protein interactions observed here might both be important in telomere biology.     

HYLAS: program for generating H curves (abstract three-dimensional representations of long DNA sequences)

The computer program HYLAS generates from a standard DNA lettersequence a three-dimensional space curve (H curve) which embodiesthe entire information content of the original nucleotide sequence.The program can display H curves either as two-dimensional (frontand side view) projections or as stereo-pair images. The curvescan be marked at specific nucleotide locations, annotated, rotatedfor observation from any viewing angle, and manipulated forconvenient side-by-side comparisons. Unlike the cumbersome lettersequences, H curves can be drastically condensed in size withoutlosing their ability to reflect the global nucleotide-distributionpattern of the entire DNA sequence. Often, biologically importantloci can be visually identified on the H curves. HYLAS is writtenin FORTRAN with separate mainframe (IBM- VM/CMS) and microcomputer(MS-DOS) versions. It uses the Tektronix-TCS library of graphicsubroutines. Received on October 24, 1988; accepted on July 15, 1989     

A computer algorithm to determine the recognition site of restriction enzymes

A new algorithm is proposed to determine the type-II restrictionendonucleases' recognition site knowing the digested DNA sequenceand fragment lengths in an actual case. The algorithm is implementedfor the Commodore 64 microcomputer. Received on January 6, 1987; accepted on June 19, 1987     

Host-specific regulation of nodulation genes in Rhizobium is mediated by a plant-signal, interacting with the nodD gene product

We have identified a nodD gene from the wide host-range Rhizobium strain MPIK3030 (termed nodD1) which is essential for nodulation on Macroptilium atropurpureum (siratro). Experiments with nodA–lacZ gene fusions demonstrate that the MPIK3030 nodD1 regulates expression of the nodABC genes. Additionally, we used nodC–lacZ fusions of Rhizobium meliloti to show that the MPIK3030 nodD1 gene induces expression of these fusions by interacting with plant factors from siratro and from the non-host Medicago sativa (alfalfa). The R. meliloti nodD genes, however, only interact with alfalfa exudate. In line with these results, no complementation of MPIK3030 nodD1 mutants could be obtained on siratro with the R. meliloti nodD genes, while the MPIK3030 nodD1 can complement nodD mutants of R. meliloti on alfalfa. Furthermore, R. meliloti transconjugants harbouring the MPIK3030 nodD1 efficiently nodulate the illegitimate host siratro. When compared with other nodD sequences, the amino acid sequence of the MPIK3030 nodD1 shows a conserved aminoterminus, whereas the carboxy-terminus of the putative gene product diverges considerably. Studies on a chimeric MPIK3030/R. meliloti nodD gene indicates that the carboxy-terminal region is responsible for the interaction with plant factor(s) and may have evolved in different rhizobia specifically to interact with plant–host factors.     

The in vitro cellular immune response to quartz

It is known that macrophages release into medium in vitro biologically active substances which modulate immune response. In recent years, increase attention has been directed towards the role of prostaglandin in macrophage function. Guinea pig splenic lymphocytes and peritoneal macrophage cultures were incubated with quartz (DQ12), Corundum and aspirin as prostaglandin inhibitor. Lymphocyte proliferation measured by 3H-thymidine incorporation, and lymphokine release by Bendixen-Soborg capillary test (MIF) were applied. EA rosetting and phagocytosis were determined for macrophage function assessment. Quartz suppressed the immune response evidenced by reduced 3H-thymidine incorporation and decreased lymphokine production. Aspirin (as a specific prostaglandin synthesis inhibitor) restored the affected by quartz immunological parameters. This observation gives support to the hypothesis that the immunological response to quartz involves macrophage prostaglandin release, presumably as a first step of lipid peroxidation.     

The inhibitory effect of succinate on radiation-enhanced mitochondrial lipid peroxidation

The degree of mitochondrial ADP/Fe/NADPH-induced lipid peroxidation was increased up to the fourth day after 9.0 Gy whole body gamma-irradiation. The lipid peroxidation inhibiting effect of succinate added to isolated mitochondria was diminished as a consequence of irradiation. The succinate, administered in vivo prior to irradiation, decreased the amount of malondialdehyde production and protected the succinate dehydrogenase enzyme against inactivation. The mean survival of succinate-pretreated animals was much longer than that of controls. The role of mitochondrial lipid peroxidation in the pathogenesis of radiation injury is discussed.     

The effect of oxidants on biomembranes and cellular metabolism

During the reductive process in the tissues, the aerobes generate a number of oxidants. Unless these oxidants are reduced, oxidative damage and cell death would occur. Oxidation of plasma membrane lipids leads to autocatalytic chain reactions which eventually alter the permeability of the cell. The role of oxidative damage in the pathophysiology of diabetic complications and ischemic reperfusion injury of myocardium, especially the changes in the channel activity which may lead to arrhythmia have been studied. Hyperglycemia activates aldose reductase which could efficiently reduce glucose to sorbitol in the presence of NADPH. Since NADPH is also aldose required by glutathione reductase for reducing oxidants, its diversion would lead to membrane lipid oxidation and permeability changes which are probably responsible for diabetic complications such as cataractogenesis, retinopathy, neuropathy etc. Antioxidants such as butylated hydroxy toluene (BHT) and also reductase inhibitors prevent or delay some of these complications. By using patch-clamp technique in isolated frog myocytes, we have shown that hydroxy radicals generated by ferrous sulfate and ascorbate as well as lipid peroxides such as t-butyl hydroperoxide facilitate the entry of Na⁺ by oxidizing Na⁺-channels. Increased intracellular Na⁺ leads to an increase in Na⁺/Ca²⁺ exchange. The increased Na⁺ concentration by itself may produce electrical disturbance which would result in arrhythmia. Increased Ca²⁺ may affect proteases and may help in the conversion of xanthine dehydrogenase to xanthine oxidase, consequently increased production of super oxide radicals. Increased membrane lipid peroxidation and other oxygen free-radical associated membrane damage in myocytes has been demonstrated.     

Lipid peroxidation in Ehrlich ascites cell mitochondria is not determined by the polyunsaturated fatty acid content of the membrane

Lipid peroxidation intensity is compared in Ehrlich Ascites Cell and in liver mitochondria, prepared from tumor bearing mice. Malondialdehyde formation is negligible in intact ascites tumour mitochondria, but it is significantly increased in permeabilised mitochondria and in isolated mitochondrial membranes. We suggest that the resistance against oxidative stress is a consequence of efficient protective mechanisms operating in the intact tumour mitochondria and the low level of polyunsaturated fatty acids under these circumstances cannot be the rate limiting factor in lipid peroxidation. Succinate, an effective inhibitor of mitochondrial lipid peroxidation in liver, cannot determine malondialdehyde formation in ascites tumour mitochondria.