Molecular insights of SAH enzyme catalysis and implication for inhibitor design

Biological transmethylation reaction is a key step in the duplication of virus life cycle, in which S-adenosylmethionine plays as the methyl donor. The product of this reactions, S-adenosylhomocysteine (AdoHcy) inhibits the transmethylation process. AdoHcy is hydrolysed to adenosine and L-homocysteine by the action of S-adenosylhomocysteine hydrolase (SAH). Thus the virus life cycle should be cut off once the action of SAH is inhibited. Our study was focussed on the discovery of potential inhibitor against SAH. We performed a similarity search in Traditional Chinese Medicine Database and retrieved 17 hits with high similarity. After that we virtually docked the 17 compounds as well as the natural substrates to the hydrolase using Autodock 3.0.1 software. Then we discussed about the mechanism of the inhibition reaction, followed by proposing the potential inhibitors by comparing best docked solutions and possible modification for the best inhibitors.     

Heterotrimeric G-protein participation in Arabidopsis pollen germination through modulation of a plasmamembrane hyperpolarization-activated Ca2+-permeable channel

The role of heterotrimeric G proteins in pollen germination and tube growth was investigated using Arabidopsis thaliana plants in which the gene (GPA) encoding the G-protein a subunit (Galpha) was null or overexpressed. Pollen germination, free cytosolic calcium concentration ([Ca(2+)](cyt)) and Ca(2+) channel activity in the plasma membrane (PM) of pollen cells were investigated. Results showed that, compared with pollen grains of the wild type (ecotype Wassilewskija, ws), in vitro germinated pollen of Galpha null mutants (gpa1-1 and gpa1-2) had lower germination percentages and shorter pollen tubes, while pollen from Galpha overexpression lines (wGalpha and cGalpha) had higher germination percentages and longer pollen tubes. Compared with ws pollen cells, [Ca(2+)](cyt) was lower in gpa1-1 and gpa1-2 and higher in wGalpha and cGalpha. In whole-cell patch clamp recordings, a hyperpolarization-activated Ca(2+)-permeable conductance was identified in the PM of pollen protoplasts. The conductance was suppressed by trivalent cations but insensitive to organic blockers; its permeability to divalent cations was Ba(2+) > Ca(2+) > Mg(2+) > Sr(2+) > Mn(2+). The activity of the Ca(2+)-permeable channel conductance was down-regulated in pollen protoplasts of gpa1-1 and gpa1-2, and up-regulated in wGalpha and cGalpha. The results suggest that Galpha may participate in pollen germination through modulation of the hyperpolarization-activated Ca(2+) channel in the PM of pollen cells.     

Proteome analysis of leaves from the resurrection plant <Emphasis Type="Italic">Boea hygrometrica</Emphasis> in response to dehydration and rehydration

Resurrection plants differ from other species in their unique ability to survive desiccation. In order to understand the mechanisms of desiccation tolerance, proteome studies were carried out using leaves of the resurrection plant Boea hygrometrica to reveal proteins that were differentially expressed in response to changes in relative water content. This opportunity was afforded by the rare ability of excised B. hygrometrica leaves to survive and resume metabolism following desiccation in a manner similar to intact plants. From a total of 223 proteins that were reproducibly detected and analyzed, 35% showed increased abundance in dehydrated leaves, 5% were induced in rehydrated leaves and 60% showed decreased or unchanged abundance in dehydrated and rehydrated leaves. Since the induction kinetics fall into clearly defined patterns, we suggest that programmed regulation of protein expression triggered by changes of water status. Fourteen dehydration responsive proteins were analyzed by mass spectrometry. Eight proteins were classified as playing a role in reactive oxygen species scavenging, photosynthesis and energy metabolism. In agreement with these findings, glutathione content and polyphenol oxidase activity were found to increase upon dehydration and rapid recovery of photosynthesis was observed.     

Repetitive deformation activates focal adhesion kinase and ERK mitogenic signals in human Caco-2 intestinal epithelial cells through Src and Rac1

Intestinal epithelial cells are subject to repetitive deformation during peristalsis and villous motility, whereas the mucosa atrophies during sepsis or ileus when such stimuli are abnormal. Such repetitive deformation stimulates intestinal epithelial proliferation via focal adhesion kinase (FAK) and extracellular signal-regulated kinases (ERK). However, the upstream mediators of these effects are unknown. We investigated whether Src and Rac1 mediate deformation-induced FAK and ERK phosphorylation and proliferation in human Caco-2 and rat IEC-6 intestinal epithelial cells. Cells cultured on collagen-I were subjected to an average 10% cyclic strain at 10 cycles/min. Cyclic strain activated Rac1 and induced Rac1 translocation to cell membranes. Mechanical strain also induced rapid sustained phosphorylation of c-Src at Tyr(418), Rac1 at Ser(71), FAK at Tyr(397) and Tyr(576), and ERK1/2 at Thr(202)/Tyr(204). The mitogenic effect of cyclic strain was blocked by inhibition of Src (PP2 or short interfering RNA) or Rac1 (NSC23766). Src or Rac1 inhibition also prevented strain-induced FAK phosphorylation at Tyr(576) and ERK phosphorylation but not FAK phosphorylation at Tyr(397). Reducing FAK using short interfering RNA blocked strain-induced mitogenicity and attenuated ERK phosphorylation but not Src or Rac1 phosphorylation. Src inhibition blocked strain-induced Rac1 phosphorylation, but Rac inhibition did not alter Src phosphorylation. Transfection of a two-tyrosine phosphorylation-deficient FAK mutant Y576F/Y577F prevented activation of cotransfected myc-ERK2 by cyclic strain. Repetitive deformation induced by peristalsis or villus motility may support the gut mucosa by a pathway involving Src, Rac1, FAK, and ERK. This pathway may present important targets for interventions to prevent mucosal atrophy during prolonged ileus or fasting.     

Different electrostatic potentials define ETGE and DLG motifs as hinge and latch in oxidative stress response

Nrf2 is the regulator of the oxidative/electrophilic stress response. Its turnover is maintained by Keap1-mediated proteasomal degradation via a two-site substrate recognition mechanism in which two Nrf2-Keap1 binding sites form a hinge and latch. The E3 ligase adaptor Keap1 recognizes Nrf2 through its conserved ETGE and DLG motifs. In this study, we examined how the ETGE and DLG motifs bind to Keap1 in a very similar fashion but with different binding affinities by comparing the crystal complex of a Keap1-DC domain-DLG peptide with that of a Keap1-DC domain-ETGE peptide. We found that these two motifs interact with the same basic surface of either Keap1-DC domain of the Keap1 homodimer. The DLG motif works to correctly position the lysines within the Nrf2 Neh2 domain for efficient ubiquitination. Together with the results from calorimetric and functional studies, we conclude that different electrostatic potentials primarily define the ETGE and DLG motifs as a hinge and latch that senses the oxidative/electrophilic stress.     

H bonding at the helix-bundle crossing controls gating in Kir potassium channels

Specific stimuli such as intracellular H+ and phosphoinositides (e.g., PIP2) gate inwardly rectifying potassium (Kir) channels by controlling the reversible transition between the closed and open states. This gating mechanism underlies many aspects of Kir channel physiology and pathophysiology; however, its structural basis is not well understood. Here, we demonstrate that H+ and PIP2 use a conserved gating mechanism defined by similar structural changes in the transmembrane (TM) helices and the selectivity filter. Our data support a model in which the gating motion of the TM helices is controlled by an intrasubunit hydrogen bond between TM1 and TM2 at the helix-bundle crossing, and we show that this defines a common gating motif in the Kir channel superfamily. Furthermore, we show that this proposed H-bonding interaction determines Kir channel pH sensitivity, pH and PIP2 gating kinetics, as well as a K+-dependent inactivation process at the selectivity filter and therefore many of the key regulatory mechanisms of Kir channel physiology.     

Studies on synthesis and anion recognition properties of (3'-nitrobenzo)[2,3-d]-(3'-nitrobenzo)[9,10-d]-1,4,8,11-tetraazacyclotetradecane-5,7,12,14-tetraone

A novel artificial receptor, (3'-nitrobenzo)[2,3-d]-(3'-nitrobenzo)[9,10-d]-1,4,8,11-tetraazacyclotetradecane-5,7,12,14-tetraone, was designed and synthesized. The interactions of this receptor with different anions were determined by UV-vis and (1)H NMR titration experiments, and their affinity constants to different anions were compared with those of other similar/different systems. The results indicated that this receptor showed a high selective and recognitive ability for F(-) among F(-), Cl(-), Br(-), I(-), AcO(-), OH(-), and H(2)PO(4)(-). Moreover, the low energy configuration of this receptor was further determined by means of theoretical investigations.     

海绵共附生微生物的研究新进展

海绵是最原始的一类后生动物,已被作为海洋活性化合物的重要来源之一,其独特的孔状结构使其成为许多海洋微生物的优良宿主。近年来国内对海绵及其共附生微生物的研究主要集中在它们产生的活性物质方面,但对海绵共附生微生物的分布、多样性及其对宿主海绵作用的研究鲜有报道,就国内外研究进展进行了综述。     

烟草抗病毒基因工程研究进展

烟草花叶病毒病是一种危害严重的世界性病害,可大幅度降低烟草的产量和品质。研究和探索防治烟草花叶病毒病的新技术、新途径已成为众多研究者普遍关注的焦点。分子生物学的发展,特别是基因工程的发展为防治该病毒病带来了希望。综述了目前普遍采用的几种烟草抗病毒基因工程策略的研究现状及评价。     

蝶类的拟态现象

拟态是一物种在形态、色型和行为上模拟另一物种,从而获得好处的行为,拟态可区分为贝次拟态、缪勒拟态和拟态集团3种类型。以蝶类为例介绍了这3种拟态的概念和实例,并论述了拟态现象的进化过程。