Construction of Mutant Genes for a Non-Toxic Verotoxin 2 Variant (VT2vp1) of Escherichia coli and Characterization of Purified Mutant Toxins

The gene encoding a Verotoxin 2 variant, VTvp1, was mutated by oligonucleotide-directed site-specific mutagenesis. Among 6 mutant toxins encoded by the mutated genes, E167Q-R170L (glutamic acid at position 167 and arginine at position 170 from N-terminus of the A subunit were replaced by glutamine and leucine, respectively) was found to have markedly decreased activities; inhibition of protein synthesis, Vero cell cytotoxicity and mouse lethality of the purified E167Q-R170L were 1/1,900, 1/125,000 and 1/2,000, respectively, of those of the purified wild-type VT2vp1. Since the antigenic property of the E167Q-R170L was demonstrated to be similar to that of the wild-type VT2vp1 by Ouchterlony double gel diffusion test and by neutralization test of Vero cell cytotoxicity of the VT2vp1, a possibility to use the mutant VT2vp1, E167Q-R170L, as a toxoid is discussed.     

三褶脉紫菀中的新二萜甙

三褶脉紫菀（ＡｓｔｅｒａｇｅｒａｔｏｉｄｅｓＴｕｒｃｚ．）系菊科多年生草本植物,遍布全国,是民间常用的中药,有清热解毒、祛痰镇咳的功效［１,２］。化学工作者们曾从其同属植物紫菀（ＡｓｔｅｒｔａｒａｒｉｃｕｓＬ．ｆ．）中分离到紫菀酮（ｓｈｉｏｎｏｎｅ）、槲皮素（ｑｕｅｒｃｅｔｉｎ）、无羁萜（ｆｒｉｅｄｅｌｉｎ）、表无羁萜（ｅｐｉｆｒｉｅｄｅｌｉｎｅｌ）、毛叶醇（ｌａｃｈｎｏｐｈｙｌｌｏｌ）、乙酸毛叶酯（ｌａｃｈｎｏｐｈｙｌｌｏｌａｃｅｔａｔｅ）、茴香醚（ａｎｅｔｈｏｌｅ）以及紫菀三萜皂甙［２—４］…     

Phylogenetic place of guinea pigs: no support of the rodent-polyphyly hypothesis from maximum-likelihood analyses of multiple protein sequences

Graur et al.'s (1991) hypothesis that the guinea pig-like rodents have an evolutionary origin within mammals that is separate from that of other rodents (the rodent-polyphyly hypothesis) was reexamined by the maximum-likelihood method for protein phylogeny, as well as by the maximum-parsimony and neighbor-joining methods. The overall evidence does not support Graur et al.'s hypothesis, which radically contradicts the traditional view of rodent monophyly. This work demonstrates that we must be careful in choosing a proper method for phylogenetic inference and that an argument based on a small data set (with respect to the length of the sequence and especially the number of species) may be unstable.      

葡萄试管苗热处理结合茎尖培养脱病毒效果的研究

对７个生食葡萄品种的试管苗热处理结合茎尖剥离培养脱除病毒的方法做了研究,结果表明,从热处理试管苗剥离的茎尖培养成活率为６１．２％,其中有１８．１％的成苗。各品种热处理试管苗剥离的茎尖分化再生能力不同：先形成愈伤组织的成苗率较低,而先分化芽的茎尖成苗率较高。这一方法对扇叶病毒和卷叶病毒的脱除率达９２％以上。     

Metabolite effectors for short-term nitrogen-dependent enhancement of phosphoenolpyruvate carboxylase activity and decrease of net sucrose synthesis in wheat leaves

It has been demonstrated previously that field pea (Pisum sativum L. cv. Express) grown in hydroponic culture on a complete nutrient solution with low NH₄⁺ concentrations (<0.5 mM) will produce a larger than normal proliferation of nodules. Peas grown in the absence of mineral N in hydroponic culture have been shown to rapidly autoregulate nodulation, forming a static nodule number by 14 to 21 days after planting. The present study further characterizes the effect of NH₄⁺ concentration in hydroponic culture on nodulation and nodule growth. Peas were grown continually for 4 weeks at NH₄⁺ concentrations that were autoregulatory (0.0 mM), stimulatory (0.2 mM) or inhibitory (1.0 mM), or peas were transferred between autoregulatory or NH₄⁺ inhibited and stimulatory solutions after 2 weeks. The peas nodulated as expected when grown under constant autoregulatory, stimulatory or inhibitory concentrations of NH₄⁺. When peas were transferred from the inhibitory (1.0 mM) to the stimulatory solution (0.2 mM) a massive proliferation of nodule primordia over the entire root system was observed within 3 days of the transfer. When they were transferred from the autoregulatory (0.0 mM) to the stimulatory (0.2 mM) solution a 10-day delay occurred before a proliferation in nodule primordia occurred at distal regions of the root system. These findings support our hypothesis that low concentrations (<1.0 mM) of NH₄⁺ in hydroponic culture cause a suppression of autoregulation in pea. In addition, the temporal and spatial differences in nodule proliferation between transfer treatments demonstrate at a whole plant level that autoregulation and NH₄⁺ inhibition suppress early nodule development via different mechanisms.     

Isolation of two novel corrinoid proteins from acetate-grown Methanosarcina barkeri.

Two corrinoid proteins with molecular sizes of 480 and 29 kDa are stably methylated by [2-14C]acetate-derived intermediates in cell extracts of aceticlastic Methanosarcina barkeri when methylreductase is inhibited by the addition of bromoethanesulfonic acid. Both 14CH3-proteins have been isolated to near homogeneity and found to be abundant soluble proteins. The larger protein possesses two subunits, of 41.4 and 30.4 kDa, in an equimolar ratio, suggesting an alpha 6 beta 6 conformation with six bound methylated corrinoids per 480-kDa molecule. The 29-kDa protein is a monomer in solution and possesses only one methylated corrinoid. All methyl groups on both proteins are photolabile, but the methylated corrinoid bound to the 29-kDa protein undergoes photolysis at a higher rate than that bound to the 480-kDa protein. The two proteins possess discrete N termini and do not appear to be forms of the same protein in equilibrium. Neither protein has an Fe4S4 cluster, and both have UV-visible spectra most similar to that of a base-on methylated corrinoid. A previously identified methylated protein, designated the unknown A 14CH3-protein, copurifies with the 480-kDa protein and has the same subunit composition. The methyl groups of both isolated 14CH3-proteins are converted to methane in cell extracts. The methylated proteins that accumulate in extracts in the presence of bromoethanesulfonic acid are demethylated by the addition of coenzyme M. Both isolated proteins are abundant novel corrinoid proteins that can methylate and be methylated by intermediates of the methanogenic pathway.     

A new computational method for cable theory problems.

We discuss a new computational procedure for solving the linear cable equation on a tree of arbitrary geometry. The method is based on a simple set of diagrammatic rules implemented using an efficient computer algorithm. Unlike most other methods, this technique is particularly useful for determining the short-time behavior of the membrane potential. Examples are presented and the convergence and accuracy of the method are discussed.     

Gut microbiome helps honeybee (Apis mellifera) resist the stress of toxic nectar plant (Bidens pilosa) exposure: Evidence for survival and immunity

Honeybee (Apis mellifera) ingestion of toxic nectar plants can threaten their health and survival. However, little is known about how to help honeybees mitigate the effects of toxic nectar plant poisoning. We exposed honeybees to different concentrations of Bidens pilosa flower extracts and found that B. pilosa exposure significantly reduced honeybee survival in a dose-dependent manner. By measuring changes in detoxification and antioxidant enzymes and the gut microbiome, we found that superoxide dismutase, glutathione-S-transferase and carboxylesterase activities were significantly activated with increasing concentrations of B. pilosa and that different concentrations of B. pilosa exposure changed the structure of the honeybee gut microbiome, causing a significant reduction in the abundance of Bartonella (p < 0.001) and an increase in Lactobacillus. Importantly, by using Germ-Free bees, we found that colonization by the gut microbes Bartonella apis and Apilactobacillus kunkeei (original classification as Lactobacillus kunkeei) significantly increased the resistance of honeybees to B. pilosa and significantly upregulated bee-associated immune genes. These results suggest that honeybee detoxification systems possess a level of resistance to the toxic nectar plant B. pilosa and that the gut microbes B. apis and A. kunkeei may augment resistance to B. pilosa stress by improving host immunity.     

TaTPP-7A positively feedback regulates grain filling and wheat grain yield through T6P-SnRK1 signalling pathway and sugar–ABA interaction

Grain size and filling are two key determinants of grain thousand-kernel weight (TKW) and crop yield, therefore they have undergone strong selection since cereal was domesticated. Genetic dissection of the two traits will improve yield potential in crops. A quantitative trait locus significantly associated with wheat grain TKW was detected on chromosome 7AS flanked by a simple sequence repeat marker of Wmc17 in Chinese wheat 262 mini-core collection by genome-wide association study. Combined with the bulked segregant RNA-sequencing (BSR-seq) analysis of an F₂ genetic segregation population with extremely different TKW traits, a candidate trehalose-6-phosphate phosphatase gene located at 135.0 Mb (CS V1.0), designated as TaTPP-7A, was identified. This gene was specifically expressed in developing grains and strongly influenced grain filling and size. Overexpression (OE) of TaTPP-7A in wheat enhanced grain TKW and wheat yield greatly. Detailed analysis revealed that OE of TaTPP-7A significantly increased the expression levels of starch synthesis- and senescence-related genes involved in abscisic acid (ABA) and ethylene pathways. Moreover, most of the sucrose metabolism and starch regulation-related genes were potentially regulated by SnRK1. In addition, TaTPP-7A is a crucial domestication- and breeding-targeted gene and it feedback regulates sucrose lysis, flux, and utilization in the grain endosperm mainly through the T6P-SnRK1 pathway and sugar–ABA interaction. Thus, we confirmed the T6P signalling pathway as the central regulatory system for sucrose allocation and source–sink interactions in wheat grains and propose that the trehalose pathway components have great potential to increase yields in cereal crops.     

Investigations of interaction mechanism and conformational variation of serum albumin affected by artemisinin and dihydroartemisinin

In this work, binding interactions of artemisinin (ART) and dihydroartemisinin (DHA) with human serum albumin (HSA) and bovine serum albumin (BSA) were investigated thoroughly to illustrate the conformational variation of serum albumin. Experimental results indicated that ART and DHA bound strongly with the site I of serum albumins via hydrogen bond (H-bond) and van der Waals force and subsequently statically quenched the intrinsic fluorescence of serum albumins through concentration-dependent manner. The quenching abilities of two drugs on the intrinsic fluorescence of HSA were much higher than the quenching abilities of two drugs on the intrinsic fluorescence of BSA. Both ART and DHA, especially DHA, caused the conformational variation of serum albumins and reduced the α-helix structure content of serum albumins. DHA with hydrophilic hydroxyl group bound with HSA more strongly, suggesting the important roles of the chemical polarity and the hydrophilicity during the binding interactions of two drugs with serum albumins. These results reveal the molecular understanding of binding interactions between ART derivatives and serum albumins, providing vital information for the future application of ART derivatives in biological and clinical areas.