4-Aminotetrolic Acid : New Conformational-restricted Analogue of γ-Aminobutyric Acid

RECENT studies strongly support a role for γ-aminobutyric acid (GABA) as an inhibitory transmitter at certain synapses in the mammalian central nervous system. Structure activity correlations of many GABA analogues implicate both the intramolecular distance between the zwitterionic centres and the rotational freedom of the molecule as important factors governing the synaptic activity of these substances¹. The following observations provide pertinent information about the active conformation(s) of GABA recognized by the receptor. (1) Muscimol, an isoxazole isolated from Amanita muscaria, seems to function as a GABA analogue as its inhibitory action on central neurones is comparable with that of GABA both in potency² and with respect to antagonism by bicuculline³. Molecular orbital calculations suggest that GABA and muscimol can assume similar conformations as zwitterions with the charged centres (N⁺ and 0^?) at least 5 and, more likely, 6 Å apart⁴. (2) The selective GABA antagonist bicuculline exhibits some degree of structural similarity with particular conformations of GABA and muscimol³. (3) X-ray crystallography indicates that GABA exists in a partially folded conformation in the solid state^5,6. (4) A model of the GABA receptor proposes that GABA adopts a folded conformation with a distance of less than 4.4 Å between the charged centres⁷. Observations (1) and (2) suggest extended conformations for GABA, while (3) and (4) suggest folded conformations.     

Metabolic Engineering for Production of β-Carotene and Lycopene in Saccharomyces cerevisiae

We have engineered a conventional yeast, Saccharomyces cerevisiae, to confer a novel biosynthetic pathway for the production of β-carotene and lycopene by introducing the bacterial carotenoid biosynthesis genes, which are individually surrounded by the promoters and terminators derived from S. cerevisiae. β-Carotene and lycopene accumulated in the cells of this yeast, which was considered to be a result of the carbon flow for the ergosterol biosynthetic pathway being partially directed to the pathway for the carotenoid production.     

Comparative study of the γ-butyrolactone and pantolactone contents in cells and musts during vinification by three Saccharomyces cerevisiae races

Summary Changes in the -butyrolactone and pantolactone contents in yeast cells and musts during fermentation and subsequent flor veil formation of Sherry wines were studied. Saccharomyces cerevisiae race cerevisiae, S. cerevisiae race bayanus and S. cerevisiae race capensis were used. During the alcoholic fermentation (first 31 days), -butyrolactone contents in musts and yeast cells were similar for the three yeast races tested. In this period, pantolactone was excreted to the must by bayanus and capensis races, and it was not detected in cerevisiae race cells. During flor veil formation (31 to 134 days), bayanus and capensis races yield higher -butyrolactone and pantolactone contents than cerevisiae race in the wines. In the final wines, pantolactone contents were always lower than those of -butyrolactone.     

Two Metabotropic γ-Aminobutyric Acid Receptors Differentially Modulate Calcium Currents in Retinal Ganglion Cells

Metabotropic γ-aminobutyric acid (GABA) receptors were studied in amphibian retinal ganglion cells using whole cell current and voltage clamp techniques. The aim was to identify the types of receptor present and their mechanisms of action and modulation. Previous results indicated that ganglion cells possess two ionotropic GABA receptors: GABA_AR and GABA_CR. This study demonstrates that they also possess two types of metabotropic GABA_B receptor: one sensitive to baclofen and another to cis-aminocrotonic acid (CACA). The effects of these selective agonists were blocked by GDP-β-S. Baclofen suppressed an ω-conotoxin–GVIA-sensitive barium current, and this action was reversed by prepulse facilitation, indicative of a direct G-protein pathway. The effect of baclofen was also partially occluded by agents that influence the protein kinase A (PKA) pathway. But the effect of PKA activation was unaffected by prepulse facilitation, indicating PKA acted through a parallel pathway. Calmodulin antagonists reduced the action of baclofen, whereas inhibitors of calmodulin phosphatase enhanced it. Antagonists of internal calcium release, such as heparin and ruthenium red, did not affect the baclofen response. Thus, the baclofen-sensitive receptor may respond to influx of calcium. The CACA-sensitive GABA receptor reduced current through dihydropyridine-sensitive channels. Sodium nitroprusside and 8-bromo-cGMP enhanced the action of CACA, indicating that a nitric oxide system can up-regulate this receptor pathway. CACA-sensitive and baclofen-sensitive GABA_B receptors reduced spike activity in ganglion cells. Overall, retinal ganglion cells possess four types of GABA receptor, two ionotropic and two metabotropic. Each has a unique electrogenic profile, providing a wide range of neural integration at the final stage of retinal information processing.     

Improvement of γ-decalactone production by stimulating the import of ricinoleic acid and suppressing the degradation of γ-decalactone in Saccharomyces cerevisiae

A number of yeast species can transform ricinoleic acid into γ-decalactone, a high-value compound with fruity aroma, through β-oxidation. This study investigated the effect of l-carnitine on γ-decalactone production by Saccharomyces cerevisiae MF013 to increase the β-oxidation rate. Results showed that l-carnitine shortened the biotransformation period by approximately 10?h and increased γ-decalactone production by 19.5%. γ-Caprolactone, γ-octalactone, and γ-dodecalactone were separately added to the medium to prevent γ-decalactone degradation by yeast cells at the end of biotransformation. γ-Octalactone competitively inhibited γ-decalactone from binding to lactonase, resulting in an 11% increase in γ-decalactone production. This research proposed an effective approach to improve the γ-decalactone production rate, shorten the biotransformation period, and suppress the γ-decalactone degradation in S. cerevisiae.     

Increasing glutathione formation by functional expression of the γ-glutamylcysteine synthetase gene in Saccharomyces cerevisiae

A recombinant plasmid, pGMF, containing a gamma-glutamylcysteine synthetase gene (GSH-I) from Saccharomyces cerevisiae, was constructed with a copper-resistance gene as the selection marker and was introduced into S. cerevisiae YSF-31. The glutathione content of the recombinant strain was 1.5-fold (13.1 mg g dry cells(-1)) of that in the host strain.     

Yeast Comparative Genetics: A New MEL15 α-Galactosidase Gene of Saccharomyces cerevisiae

To supplement the earlier identified European family of the highly homologous -galactosidase MEL1–MEL11genes and the African family of the divergent MEL12–MEL14 genes, a new MEL15gene was found in several Saccharomyces cerevisiae strains isolated from maize dough in Ghana. Southern blotting and restriction enzyme analysis assigned the MEL15 gene to the African family and mapped it to chromosomes IV/XII, which migrate together in electrophoresis. Tetrad analysis ruled out the MEL15 location in the left arm of chromosome IV or the right arm of chromosome XII, which respectively contain the known MEL5 and MEL10 genes.     

Production of γ-Aminobutyric Acid and Its Supplementary Role in the TCA Cycle in Rice (Oryza sativa L.) Seedlings

Journal of Plant Growth Regulation - Germination of rice (Oryza sativa L.) results in rapid accumulation of γ-aminobutyric acid (GABA), but its precise role in the process remains unclear. In...     

Perceptions and Patterns of Human–elephant Conflict in Old and New Settlements in Sri Lanka: Insights for Mitigation and Management

Human–elephant conflict poses a major threat to elephants in many parts of Asia, including Sri Lanka. We studied human–elephant conflict in two areas with contrasting scenarios of landuse and conflict, Kahalle and Yala. Kahalle was developed and settled under the Mahaweli irrigation project and the main agricultural practice was irrigated agriculture, with two annual growing seasons. The area was a mosaic of settlements, agriculture, and small forest patches with ill defined human- and elephant-use areas. Elephants ranged within the habitat mosaic year round, occupying remnant forest patches and raiding adjacent crops at night. In contrast, Yala was dominated by a large protected area complex, and the main agricultural methods were slash-and-burn agriculture and rain-fed paddy cultivation. Human- and elephant-use areas were well defined and segregated. The protected area provided elephants with a refuge and food during the rainy season, when the single annual crop was grown. During the dry season, elephants moved into slash-and-burn areas and utilized leftover crops and pioneer vegetation in fallow fields. The landuse pattern and agricultural practices in Yala facilitated co-existence, whereas that in Kahalle led to year round conflict. We suggest that areas managed according to traditional landuse practices should be part of an elephant conservation strategy, where people and elephants have to share resources.     

TGFβ Activated Kinase-1: New Insights into the Diverse Roles of TAK1 in Development and Immunity

A number of recent publications have examined the role of TAK1 in model systems ranging from fly to mouse. Rather than fit into a clearly defined linear molecular pathway, TAK1 seems to act in a signaling nexus that responds to a variety of upstream signals, including inflammatory molecules and developmental cues. TAK1 then influences a number of downstream processes ranging from innate immune responses to patterning and differentiation via JNK, NFκB, and TCFβ-catenin signaling. These differences in function are not simply a matter of cell type. For example, NFκB signaling in a particular cell may or may not require TAK1 depending on the nature of the activating signal. Interestingly, the multi-task functionality of TAK1 is conserved between vertebrate and invertebrate species. Studies of TAK1 in multiple experimental systems is likely to reveal more roles for this kinase and also elucidate mechanisms by which other signaling molecules fulfill diverse signaling roles. Here we provide an overview of the data concerning TAK1 from its discovery to more recent findings and provide a synthesis of the conclusions that have arisen from the multiple model systems and experimental approaches.     

New insights into the role of mast cells in autoimmunity: Evidence for a common mechanism of action?

Mast cells are classically considered innate immune cells that act as first responders in many microbial infections and have long been appreciated as potent contributors to allergic reactions. However, recent advances in the realm of autoimmunity have made it clear that these cells are also involved in the pathogenic responses that exacerbate disease. In the murine models of multiple sclerosis, rheumatoid arthritis and bullous pemphigoid, both the pathogenic role of mast cells and some of their mechanisms of action are shared. Similar to their role in infection and a subset of allergic responses, mast cells are required for the efficient recruitment of neutrophils to sites of inflammation. Although this mast cell-dependent neutrophil response is protective in infection settings, it is postulated that neutrophils promote local vascular permeability and facilitate the entry of inflammatory cells that enhance tissue destruction at target sites. However, there is still much to learn. There is little information regarding mechanisms of mast cell activation in disease. Nor is it known how many mast cell-derived mediators are relevant and whether interactions with other cells are implicated in these diseases including T cells, B cells and astrocytes. Here we review the current state of knowledge about mast cells in autoimmune disease. We also discuss findings regarding newly discovered mast cell actions and factors that modulate mast cell function. We speculate that much of this new information will ultimately contribute to a greater understanding of the full range of mast cell actions in autoimmunity. This article is part of a Special Issue entitled: Mast cells in inflammation.     

Evidence for conserved function of γ-glutamyltranspeptidase in Helicobacter genus

The confounding consequences of Helicobacter bilis infection in experimental mice populations are well recognized, but the role of this bacterium in human diseases is less known. Limited data are available on virulence determinants of this species. In Helicobacter pylori, γ-glutamyltranspeptidase (γGT) contributes to the colonization of the gastric mucosa and to the pathogenesis of peptic ulcer. The role of γGT in H. bilis infections remains unknown. The annotated genome sequence of H. bilis revealed two putative ggt genes and our aim was to characterize these H. bilis γGT paralogues. We performed a phylogenetic analysis to understand the evolution of Helicobacter γGTs and to predict functional activities of these two genes. In addition, both copies of H. bilis γGTs were expressed as recombinant proteins and their biochemical characteristics were analysed. Functional complementation of Esherichia coli deficient in γGT activity and deletion of γGT in H. bilis were performed. Finally, the inhibitory effect of T-cell and gastric cell proliferation by H. bilis γGT was assessed. Our results indicated that one gene is responsible for γGT activity, while the other showed no γGT activity due to lack of autoprocessing. Although both H. bilis and H. pylori γGTs exhibited a similar affinity to L-Glutamine and γ-Glutamyl-p-nitroanilide, the H. bilis γGT was significantly less active. Nevertheless, H. bilis γGT inhibited T-cell proliferation at a similar level to that observed for H. pylori. Finally, we showed a similar suppressive influence of both H. bilis and H. pylori γGTs on AGS cell proliferation mediated by an apoptosis-independent mechanism. Our data suggest a conserved function of γGT in the Helicobacter genus. Since γGT is present only in a few enterohepatic Helicobacter species, its expression appears not to be essential for colonization of the lower gastrointestinal tract, but it could provide metabolic advantages in colonization capability of different niches.     

Secretory expression of β-mannanase in Saccharomyces cerevisiae and its high efficiency for hydrolysis of mannans to mannooligosaccharides

Applied Microbiology and Biotechnology - Degradation of mannans is a key process in the production of foods and prebiotics. β-Mannanase is the key enzyme that hydrolyzes...     

Manganese toxicity towards Saccharomyces cerevisiae : Dependence on intracellular and extracellular magnesium concentrations

Inhibition of the growth of Saccharomyces cerevisiae was evident at concentrations of 0.5 mM Mn²⁺ or higher, but a tolerance to lower Mn²⁺ concentrations was observed. The inhibitory effects of 2.0 mM Mn²⁺ were eliminated by supplementing the medium with excess Mg²⁺ (10 mM), whereas addition of excess Ca²⁺ and K⁺ had negligible effect on Mn²⁺ toxicity. Growth inhibition by Mn²⁺, in the absence of a Mg²⁺ supplement, was attributed to Mn²⁺ accumulation to toxic intracellular levels. Mn levels in S. cerevisiae grown in Mg²⁺-supplemented medium were severalfold lower than those of cells growing in unsupplemented medium. Mn²⁺ toxicity was also influenced by intracellular Mg, as Mn²⁺ toxicity was found to be more closely correlated with the cellular Mg:Mn ratio than with cellular Mn levels alone. Cells with low intracellular levels of Mg were more susceptible to Mn²⁺ toxicity than cells with high cellular Mg, even when sequestered Mn²⁺ levels were similar. A critical Mg:Mn ratio of 2.0 was identified below which Mn²⁺ toxicity became acute. The results demonstrate the importance of intracellular and extracellular competitive interactions in determining the toxicity of Mn²⁺. Received: 18 June 1997 / Received last revision: 10 January 1998 / Accepted: 24 January 1998