(alphaMe)Nva: stereoselective syntheses and preferred conformations of selected model peptides.

Using different stereoselective chemical and chemoenzymatic approaches we synthesized the chiral, Calpha-methylated alpha-amino acid L-(alphaMe)Nva with a short, linear side-chain. A set of terminally protected model peptides to the pentamer level containing either (alphaMe)Nva or Nva in combination with Ala and/or Aib was prepared using solution methods and characterized fully. Two (alphaMe)Nva peptides were also synthesized using side-chain hydrogenation of the corresponding Calpha-methyl, Calpha-allylglycine (Mag) peptides. A detailed solution and crystal-state conformational analysis based on FT-IR absorption, 1H NMR and X-ray diffraction techniques allowed us to define that: (i) (alphaMe)Nva is an effective beta-turn and 3(10)-helix former; and (ii) the relationship between (alphaMe)Nva chirality and the screw sense of the turn/helix formed is that typical of protein amino acids, i.e. L-(alphaMe)Nva induces the preferential formation of right-handed folded structures. In more general terms, this study reinforced previous conclusions that peptides based on alpha-amino acids with a Calpha-methyl substituent and a Calpha-linear alkyl substituent are characterized by a strong tendency to fold into turn and helical structures.     

Effect of Rhizobium–Azospirillum coinoculation on nitrogen fixation and yield of two contrasting Phaseolus vulgaris L. genotypes cultivated across different environments in Cuba

Azospirillum spp. have shown potential to enhance nodulation and plant growth of legumes when coinoculated with Rhizobium. The effect of Azospirillum on the Rhizobium-legume symbiosis is, however, dependent on the host genotype used. Previous greenhouse experiments identified two genotypes of common bean (Phaseolus vulgaris L.), BAT477 and DOR364, contrasting in nodulation response to Azospirillum when coinoculated with Rhizobium. Genetic analysis revealed a genetic basis (Quantitative Trait Loci) on the bean genome related to the differential responsiveness to Azospirillum between the two bean genotypes. In this study, on-station and on-farm field experiments in different regions in Cuba were conducted to evaluate the agronomic relevance of the differences in response to Azospirillum–Rhizobium coinoculation between the two genotypes BAT477 and DOR364. It was observed that Azospirillum–Rhizobium coinoculation as compared to single Rhizobium inoculation increased the amount of fixed nitrogen and the yield of DOR364 across all sites. For BAT477, on the contrary, a negative effect of Azospirillum–Rhizobium coinoculation on yield and nitrogen fixation was observed on most of the sites as compared to single Rhizobium inoculation. The modified stability regression equations resulting from this study may contribute to predict how a combination of genotype and inoculum will perform at a certain environmental setting. This study highlights the importance of genotype × inocula interactions in agricultural outputs and establishes a link between greenhouse phenotype, genetic background and performance in the field.     

Effect of tillage and farming system upon VAM fungus populations and mycorrhizas and nutrient uptake of maize

Low-input agricultural systems that do not rely on fertilizers may be more dependent on vesicular-arbuscular mycorrhizal [VAM] fungi than conventionally managed systems. We studied populations of spores of VAM fungi, mycorrhiza formation and nutrient utilization of maize (Zea mays L.) grown in moldboard plowed, chisel-disked or no-tilled soil under conventional and low-input agricultural systems. Maize shoots and roots were collected at four growth stages. Soils under low-input management had higher VAM fungus spore populations than soils under conventional management. Spore populations and colonization of maize roots by VAM fungi were higher in no-tilled than in moldboard plowed or chisel-disked soil. The inoculum potential of soil collected in the autumn was greater for no-till and chisel-disked soils than for moldboard plowed soils and greater for low-input than conventionally farmed soil. The effects of tillage and farming system on N uptake and utilization varied with growth stage of the maize plants. The effect of farming system on P use efficiency was significant at the vegetative stages only, with higher efficiencies in plants under low-input management. The effect of tillage was consistent through all growth stages, with higher P use efficiencies in plants under moldboard plow and chisel-disk than under no-till. Plants grown in no-tilled soils had the highest shoot P concentrations throughout the experiment. This benefit of enhanced VAM fungus colonization, particularly in the low-input system in the absence of effective weed control and with likely lower soil temperatures, did not translate into enhanced growth and yield.     

The mechanisms and costs of physiological and toxicological acclimation to waterborne silver in juvenile rainbow trout (Oncorhynchus mykiss)

Juvenile rainbow trout were exposed to 0, 0.1, 1, 3, and 5 micro g/l silver (Ag, as AgNO3) for 23 days. Specific growth rate, cumulative food consumption, food-conversion efficiency, and critical swimming speed (U(crit)) were significantly reduced during 5 micro g/l Ag exposure, demonstrating a physiological cost of silver acclimation. Only the 5 microg/l Ag treatment had significant cumulative mortality (5.2%). Fish were most susceptible to silver on days 5 and 15. Exposure to 5 microg/l Ag significantly lowered plasma Na+ and Cl- on days 5 and 10, but plasma ion concentration recovered thereafter. Unidirectional Na+ uptake and gill Na/K-ATPase activity were significantly inhibited by 3 and 5 microg/l Ag exposure. Na+ uptake was inhibited by 3 micro g/l Ag at day 5 alone, whereas the effects at the highest Ag exposure persisted until day 15. Gill Na/K-ATPase was inhibited on day 5 in both the 3 and 5 microg/l Ag treatments but increased to approx. 1.5 times of control levels by day 23. Only the 3 and 5 microg/l Ag treatments produced toxicological acclimation (at least twofold elevations in 168-h LC50 values in fish subsampled on days 15 and 23). We conclude that physiological acclimation results from compensatory changes in Na+ transport at the gills, and that these changes may eventually lead to toxicological acclimation.     

How do G-protein-coupled receptors work? The case of metabotropic glutamate and GABA receptors

G-protein coupled receptors (GPCRs) represent the largest membrane proteins family in animal genomes. Being the receptors for most hormones and neurotransmitters, these proteins play a central role in intercellular communication. GPCRs can be classified into several groups based on the sequence similarity of their common structural feature: the heptahelical domain. The metabotropic receptors for the main neurotransmitters glutamate and gamma-aminobutyric acid (GABA) belong to the class III of GPCRs, together with others receptors for Ca2+, for sweet and amino acid taste compounds and for some pheromones, as well as for odorants in fish. Besides their transmembrane heptahelical domain responsible for G-protein activation, most of class III receptors possess a large extracellular domain responsible for ligand recognition. The recent resolution of the structure of this binding domain of one of these receptors, the mGlu1 receptor, together with the recent demonstration that these receptors are dimers, revealed an original mechanism of activation for these GPCRs. Such data open new possibilities to develop drugs aimed at modulating these receptors, and raised a number of interesting questions on the activation mechanism of other GPCRs.     

The activation mechanism of class-C G-protein coupled receptors

Class-C G-protein coupled receptors (GPCRs) represent a distant group among the large family of GPCRs. This class includes the receptors for the main neurotransmitters, glutamate and gamma-aminobutyric acid (GABA), and the receptors for Ca(2+), some taste and pheromone molecules, as well as some orphan receptors. Like any other GPCRs, class-C receptors possess a heptahelical domain (HD) involved in heterotrimeric G-protein activation, but most of them also have a large extracellular domain (ECD) responsible for agonist recognition and binding. In addition, it is now well accepted that these receptors are dimers, either homo or heterodimers. This complex architecture raises a number of important questions. Here we will discuss our view of how agonist binding within the large ECD triggers the necessary change of conformation, or stabilize a specific conformation, of the heptahelical domain leading to G-protein activation. How ligands acting within the heptahelical domain can change the properties of these complex macromolecules.     

Mapping the agonist-binding site of GABAB type 1 subunit sheds light on the activation process of GABAB receptors

The gamma-amino-n-butyric acid type B (GABA(B)) receptor is composed of two subunits, GABA(B)1 and GABA(B)2, belonging to the family 3 heptahelix receptors. These proteins possess two domains, a seven transmembrane core and an extracellular domain containing the agonist binding site. This binding domain is likely to fold like bacterial periplasmic binding proteins that are constituted of two lobes that close upon ligand binding. Here, using molecular modeling and site-directed mutagenesis, we have identified residues in the GABA(B)1 subunit that are critical for agonist binding and activation of the heteromeric receptor. Our data suggest that two residues (Ser(246) and Asp(471)) located within lobe I form H bonds and a salt bridge with carboxylic and amino groups of GABA, respectively, demonstrating the pivotal role of lobe I in agonist binding. Interestingly, our data also suggest that a residue within lobe II (Tyr(366)) interacts with the agonists in a closed form model of the binding domain, and its mutation into Ala converts the agonist baclofen into an antagonist. These data demonstrate the pivotal role played by the GABA(B)1 subunit in the activation of the heteromeric GABA(B) receptor and are consistent with the idea that a closed state of the binding domain of family 3 receptors is required for their activation.     

Inhibition of reproductive maturation and somatic growth of Fischer 344 rats by photoperiods shorter than L14:D10 and by gradually decreasing photoperiod

Photoperiod is the major regulator of reproduction in temperate-zone mammals. Laboratory rats are generally considered to be nonphotoresponsive, but young male Fischer 344 (F344) rats have a uniquely robust response to short photoperiods of 8 h of light. Rats transferred at weaning from a photoperiod of 16 h to photoperiods of < 14 h of light slowed in both reproductive development and somatic growth rate. Those in photoperiods < 13 h of light underwent the strongest responses. The critical photoperiod of F344 rats can be defined as 13.5 h of light, but photoperiods of 相似文献   

Three-dimensional model of the extracellular domain of the type 4a metabotropic glutamate receptor: new insights into the activation process

Metabotropic glutamate receptors (mGluRs) belong to the family 3 of G-protein-coupled receptors. On these proteins, agonist binding on the extracellular domain leads to conformational changes in the 7-transmembrane domains required for G-protein activation. To elucidate the structural features that might be responsible for such an activation mechanism, we have generated models of the amino terminal domain (ATD) of type 4 mGluR (mGlu4R). The fold recognition search allowed the identification of three hits with a low sequence identity, but with high secondary structure conservation: leucine isoleucine valine-binding protein (LIVBP) and leucine-binding protein (LBP) as already known, and acetamide-binding protein (AmiC). These proteins are characterized by a bilobate structure in an open state for LIVBP/LBP and a closed state for AmiC, with ligand binding in the cleft. Models for both open and closed forms of mGlu4R ATD have been generated. ACPT-I (1-aminocyclopentane 1,3,4-tricarboxylic acid), a selective agonist, has been docked in the two models. In the open form, ACPT-I is only bound to lobe I through interactions with Lys74, Arg78, Ser159, and Thr182. In the closed form, ACPT-I is trapped between both lobes with additional binding to Tyr230, Asp312, Ser313, and Lys317 from lobe II. These results support the hypothesis that mGluR agonists bind a closed form of the ATDs, suggesting that such a conformation of the binding domain corresponds to the active conformation.