DNA cytosine methylation and heat-induced deamination

The heat-induced conversion of 5-methylcytosine (m⁵C) residues to thymine residues and of cytosine to uracil residues in single-stranded DNA was studied. The calculated rates for deamination at 37°C and pH 7.4 were 9.5×10^–10 and 2.1×10^–10 sec^–1, respectively. N⁴-Methyldeoxycytidine, which is in the DNA of certain thermophilic bacteria, was more heat-resistant than was deoxycytidine and much more than was 5-methyldeoxycytidine. Thermophilic bacteria which contain N⁴-methylcytosine rather than m⁵C in their genomes may thereby largely avoid heat-induced mutation due to deamination, which is incurred by the many organisms that contain m⁵C in their DNA.     

Effects of nicotine and dietary salt on a learned blood pressure response in Dahl-S rats

We examined the effects of chronic nicotine exposure and dietary salt on the arterial blood pressure (BP) changes learned in response to an acute behavioral stress in the Dahl salt-sensitive rat. Four groups were tested: low salt + vehicle; low salt + nicotine; high salt + vehicle; and high salt + nicotine. Rats were fed a low-salt (0.08% NaCl) or a high-salt (8% NaCl) diet for 4 wk; 2.4 mg. kg(-1). day(-1) nicotine or vehicle was given via an implanted osmotic minipump for the last 2 wk. All rats were conditioned by following one tone (CS+) with a 0.5-s tail shock; another tone (CS-) was never followed by shock. CS+ in low salt + vehicle and high salt + vehicle-treated rats evoked an initial arterial BP increase (C(1)), a component of the startle response, and an ensuing, smaller, but more sustained, pressor response (C(2)), which is acquired with training. In these rats, both C(1) and C(2) evoked by CS- were significantly smaller than those to CS+, demonstrating that these groups discriminated between the two tests. Conversely, although the low salt + nicotine-treated rats had both the C(1) and C(2) components of the conditional arterial pressure response, they did not discriminate between CS+ and CS-. Finally, the high salt + nicotine group failed to both discriminate between tones and acquire (i.e., learn) the C(2) response. The unconditional response to shock did not differ between groups. We conclude that combined exposure to high salt and to nicotine inhibits the salt-sensitive animal's acquisition of a learned conditional BP response, perhaps because nicotine acts preferentially on those central processes required for associative learning versus those involved in orientating to external stimuli.     

The Blomia tropicalis allergens

Blomia tropicalis allergens are the most important mite allergens in tropical regions. Most of them only have 30-40% sequence identity with their Dermatophagoides counterparts and they share low IgE cross reactivity and exhibit different immunobiology. Unlike the pyroglyphid counterparts, Blo t 5 is the major allergen whereas Blo t 1 only has modest allergenicity.     

Aldehyde dehydrogenase. Maintaining critical active site geometry at motif 8 in the class 3 enzyme.

Alignment of all known, diverse members of the aldehyde dehydrogenase (ALDH) extended family revealed only two strictly conserved, nonglycine residues, a glutamate and a phenylalanine residue. Both occur in one of the highly conserved 'motif' segments and both occupy strategic locations in the tertiary structure at the bottom of the catalytic funnel. In class 3 ALDH, these are Glu333 and Phe335. In addition, Asp247, which is not highly conserved but is characteristic of class 3 ALDHs, hydrogen bonds the main chain between Glu333 and Phe335. These three residues were mutated conservatively. Michaelis constants determined for both NAD/propanal and NADP/benzaldehyde substrate pairs show all three residues to be crucial to effective catalysis, and suggest that the hydrogen bond to Asp247 is a key element in maintaining precise geometry of key elements at the active site.     

Proliferation‐ and migration‐enhancing effects of ginseng and ginsenoside Rg1 through IGF‐I‐ and FGF‐2‐signaling pathways on RSC96 Schwann cells

The aim of the present study is to evaluate the proliferation‐ and migration‐enhancing effects of ginseng and its component, ginsenoside (Rg1) on RSC96 Schwann cells. We investigated the molecular signaling pathways, which include: (1) survival signaling, IGFs‐IGFIR‐Akt‐Bcl2 and proliferative signaling, cell cycle factors and mitogen‐activated protein kinase (MAPK) pathways, (2) migrating and anti‐scar signaling, FGF‐2‐uPA‐MMPs.We treated RSC96 cells with different concentrations (100, 200, 300, 400, 500 µg ml^?1) of ginseng and its constituent, Rg1 (5, 10, 15, 20, 25 µg ml^?1). We observed a proliferative effect in a dose‐dependent manner by PCNA western blotting assay, MTT assay, and wound healing test. Furthermore, we also found in the results of western blotting assay, ginseng and Rg1 enhance protein expression of IGF‐I pathway regulators, cell cycle controlling proteins, and MAPK signaling pathways to promote the cell proliferation. In addition, ginseng and Rg1 also stimulated the FGF‐2‐uPA‐MMP 9 migrating pathway to enhance the migration of RSC96 Schwann cells. Using MAPK chemical inhibitors, U0126, SB203580, and SP600125, the proliferative effects of ginseng and Rg1 on RSC96 cells were identified to be MAPK signaling‐dependent. On the basis of the results, applying appropriate doses of ginseng and Rg1 with biomedical materials would be a potential approach for enhancing neuron regeneration. Copyright © 2009 John Wiley & Sons, Ltd.     

Lactate dehydrogenase genes of caiman and Chinese soft-shelled turtle, with emphasis on the molecular phylogenetics and evolution of reptiles.

L-Lactate dehydrogenase (LDH) cDNAs encoding for LDH-A(4) (muscle) and LDH-B(4) (heart) isozymes from caiman (Caiman crocodilus apaporiensis) belonging to the order Crocodilia and Chinese soft-shelled turtle (Pelodiscus sinensis) belonging to the order Chelonia were sequenced. The phylogenetic relationships of the newly determined cDNA and their deduced protein sequences, as well as the previously published sequences of vertebrate LDH isozymes, were analyzed by various phylogenetic tree construction methods. These results indicated that Chelonia is indeed more closely related to Crocodilia. The divergent times between caiman and alligator, turtle and soft-shelled turtle, and Chelonia and Crocodilia were estimated to be approximately 36, 100 and 177 million years, respectively.     

Genomic organization of the chicken CD8 locus reveals a novel family of immunoreceptor genes

The genomic organization of the chicken CD8alpha gene was investigated to determine the basis of its polymorphism. Contiguous to the CD8alpha gene we identified multiple DNA blocks possessing sequences homologous to CD8alpha. Gene conversions and recombination over evolutionary time among CD8alpha and these CD8alpha homologous genes seem to account for the observed polymorphism. Furthermore, these CD8alpha-like DNAs encode a distinct multigene family of immunoreceptors that have a charged or polar residue in place of the interspecies-conserved CD8alpha transmembrane proline residue and a short cytoplasmic tail nonhomologous to CD8alpha. The identification of this novel multigene family with an organization reminiscent of human killer Ig-like receptors raises compelling questions on their evolutionary relationship among immunoreceptors.     

Structural basis of mercury- and zinc-conjugated complexes as SARS-CoV 3C-like protease inhibitors

Five active metal-conjugated inhibitors (PMA, TDT, EPDTC, JMF1586 and JMF1600) bound with the 3C-like protease of severe acute respiratory syndrome (SARS)-associated coronavirus were analyzed crystallographically. The complex structures reveal two major inhibition modes: Hg(2+)-PMA is coordinated to C(44), M(49) and Y(54) with a square planar geometry at the S3 pocket, whereas each Zn(2+) of the four zinc-inhibitors is tetrahedrally coordinated to the H(41)-C(145) catalytic dyad. For anti-SARS drug design, this Zn(2+)-centered coordination pattern would serve as a starting platform for inhibitor optimization.