A molecular dynamics study of a model built Pro-36-Gly mutant derived from the potato carboxypeptidase A inhibitor protein

A 120ps molecular dynamics (MD) trajectory was calculated and analyzed for a putative Pro-36-Gly mutant of the potato carboxypeptidase A (CPA) protein inhibitor (PCIm). The mutant protein's fold shows a large degree of stability, judged from its low alpha-carbon r.m.s. deviation from the X-ray structure of the wild type PCI (PCIw). The N-terminal tail of PCIm differs slightly less from the X-ray structure than it does in PCIw, while the mutant's C-terminal tail (the primary contact site with CPA) and residues 13-17 present deviations as they approach each other. Differences in fluctuation pattern exist between PCIm and PCIw in residues 2-4 (the N-terminal tail), 13-17, 22-23, 28-31 (the secondary contact site with CPA) and 37-38 (the C-terminal tail); the latter region is rigidified in PCIm. Results show that the MD method is able to sense local perturbative effects produced by amino acid substitutions in flexible regions of protein molecules. The simulation suggests that the conformation of the C-terminal tail is less favorable for interaction with the target protein in the mutant than it is in the wild type protein. The Pro-36-Gly mutant is predicted to be a less potent inhibitor.     

Physical and enzymatic properties of a class III isozyme of human liver alcohol dehydrogenase: chi-ADH

chi-Alcohol dehydrogenase (chi-ADH), a class III isozyme characterized by its anodic electrophoretic mobility and lack of inhibition by 4-methylpyrazole, has been isolated from human liver and purified to homogeneity in a reducing medium. chi-ADH resembles other human liver ADH isozymes of classes I and II with respect to its molecular weight, dimeric structure, stoichiometry of zinc and NADH binding, and pH optima for the oxidation of alcohols. This homodimer exhibits subtle differences in its absorption spectrum and amino acid composition relative to those of other human isozymes but differs markedly from their specificity toward alcohols and aldehydes. chi-ADH oxidizes ethanol very poorly. The reaction is bimolecular, and an apparent Km cannot be discerned up to 2.3 M ethanol. The enzyme is inactive toward methanol, ethylene glycol, digitoxigenin, digoxigenin, and gitoxigenin , but alcohols with carbon chain lengths greater than four are oxidized rapidly with Km values decreasing with increasing carbon chain length. Taken jointly, the composition, structure, and enzymatic properties of the ADH isozymes purified and studied so far strongly imply that their metabolic roles, yet to be discovered, will give a new perspective to ethanol metabolism and pathology.     

Control of Redox Balance by the Stringent Response Regulatory Protein Promotes Antioxidant Defenses of Salmonella

We report herein a critical role for the stringent response regulatory DnaK suppressor protein (DksA) in the coordination of antioxidant defenses. DksA helps fine-tune the expression of glutathione biosynthetic genes and discrete steps in the pentose phosphate pathway and tricarboxylic acid cycle that are associated with the generation of reducing power. Control of NAD(P)H/NAD(P)⁺ redox balance by DksA fuels downstream antioxidant enzymatic systems in nutritionally starving Salmonella. Conditional expression of the glucose-6-phosphate dehydrogenase-encoding gene zwf, shown here to be under DksA control, increases both the NADPH pool and antioxidant defenses of dksA mutant Salmonella. The DksA-mediated coordination of redox balance boosts the antioxidant defenses of stationary phase bacteria. Not only does DksA increase resistance of Salmonella against hydrogen peroxide (H₂O₂), but it also promotes fitness of this intracellular pathogen when exposed to oxyradicals produced by the NADPH phagocyte oxidase in an acute model of infection. Given the role of DksA in the adjustment of gene expression in most bacteria undergoing nutritional deprivation, our findings raise the possibility that the control of central metabolic pathways by this regulatory protein maintains redox homeostasis essential for antioxidant defenses in phylogenetically diverse bacterial species.     

Glucosinolate contents in maca (Lepidium peruvianum Chacón) seeds, sprouts, mature plants and several derived commercial products

Several products derived from processed maca hypocotyls (Lepidium peruvianum Chacón, previously known asL. meyenii Walp.) were surveyed for glucosinolate content and quantified by HPLC analysis. These included pills, capsules, flour, liquor, tonic and mayonnaise. Different plant organs such as fresh hypocotyls and leaves, seeds, dry hypocotyls, and sprouts were also included in the survey. The most abundant glucosinolates detected in fresh and dry hypocotyls and leaves were the aromatic glucosinolates, benzylglucosinolate (glucotropaeolin) and p-methoxybenzylglucosinolate. Maca seeds and sprouts differed in profile from hypocotyls and leaves due to the modification of benzylglucosinolate. No glucosinolates were detected in liquor and tonic, while mayonnaise had only trace amounts of those glucosinolates. It had instead allylglucosinolate (sinigrin), which is an aliphatic glucosinolate. The pills, capsules and flour had the same glucosinolates as those observed in hypocotyls, but in variable amounts. The richest sources of glucosinolates were seeds, fresh hypocotyls and sprouts, in that order.     

Capsular polysaccharide expressed by Pasteurella piscicida grown in vitro

Abstract Pasteurella piscicida grown in a glucose-rich medium produces a capsule that can be see under light and electron microscopy. The capsular polysaccharide was purified and characterized by chemical and HPLC analysis. The polymer has the composition glucose/mannose/ N -acetylgalactosamine/galacturonic acid/acetic acid in the molar ratios of approximately 2.5:1.3:0.5:0.4:2.5. The polysaccharide was immunogenic in rabbits and did not cross-react with antibodies against the O-antigen lipopolysaccharide.     

5-Hydroxytryptamine stimulates Rb efflux from pancreatic β-cells

The effects of 5-hydroxytryptamine and 5-hydroxytryptophan on ⁸⁶Rb⁺ efflux from prelabelled ob/ob-mouse islets were studied to better understand the cellular mechanisms underlying the effects of 5-hydroxytryptamine and 5-hydroxytryptophan on insulin release. 5-Hydroxytryptophan (4 mM) had no effect on ⁸⁶Rb⁺ efflux either at a low (3 mM) or at a high (20 mM) d-glucose concentration, whereas 5-hydroxytryptamine (4 mM) stimulated ⁸⁶Rb⁺ efflux at both glucose concentrations. These results indicate that 5-hydroxytryptamine may reduce glucose-induced insulin release by inhibiting early steps in the β-cell stimulus-secretion coupling.     

Does the VP1 gene of foot-and-mouth disease virus behave as a molecular clock?

Summary We have carried out a phylogenetic study of the evolution of the VP1 gene sequence from different serological types and subtypes of foot-and-mouth disease virus (FMDV). The maximum-likelihood method developed by Hasegawa and co-workers (Hasegawa et al. 1985) for the estimation of evolutionary parameters and branching dates has been used to decide between alternative models of evolution: constant versus variable rates. The results obtained indicate that a constant rate model, i.e., a molecular clock, seems to be the most plausible one. However, additional information suggests the possibility that the appearance of serotype CS has been accompanied by an episode of rapid evolution (Villaverde et al. 1991). We discuss the possibility that this evolution of RNA viruses was due to episodic positive Darwinian selection, which would have helped the new variant to escape the immunogenic pressure from the hosts. Offprint requests to: A. Moya     

Behavioural modification of a social spider by a parasitoid wasp

1. Manipulation of host behaviour by parasitoids has long captured the imagination of ecologists. Parasitoid wasps in the Polysphincta group of genera develop as external parasitoids of spiders. 2. In the present study, the previously undescribed interaction between a Zatypota sp. wasp (Ichneumonidae) and a social spider Anelosimus eximius (Theridiidae) is described. The larva of this Zatypota wasp is found to induce its host to disperse from their communal web and build an entirely enclosed web consisting of densely spun silk. 3. The wasp is observed to target primarily immature A. eximius individuals, with 37.5–44% of nests in a given area being parasitised. Of those nests, approximately 1.3–2.0% of individuals are hosts to the parasitoid larvae. Larger spider colonies had a significantly higher probability of harbouring parasitoids. 4. This interaction results in unusual behaviours for A. eximius induced by the parasitoid: (i) leaving the protection of the social nest and (ii) building a unique, altered web that it would not otherwise build. It is suggested that the wasp may be tapping into ancestral dispersal behaviours in its host and that a social species provides this wasp an evolutionary advantage by allowing a stable host source.