The presence of free d-aspartate in marine macroalgae is restricted to the Sargassaceae family

The presence of d-aspartate (d-Asp), a biologically rare amino acid, was evaluated in 38 species of marine macroalgae (seaweeds). Despite the ubiquitous presence of free l-Asp, free d-Asp was detected in only 5 species belonging to the Sargassaceae family of class Phaeophyceae (brown algae) but not in any species of the phyla Chlorophyta (green algae) and Rhodophyta (red algae). All other members of Phaeophyceae, including 3 species classified into the section Teretia of Sargassaceae did not contain d-Asp. These results indicate that the presence of free d-Asp in marine macroalgae is restricted only to the Sargassaceae family, excluding the species in the section Teretia.     

Ontogenetic appearance of somatostatin-containing nerve terminals in the median eminence of rats

Summary The development of immunoreactive (ir) somatostatin-containing nerve terminals in the rat median eminence (ME) has been examined electron-microscopically. Nerve fibers containing ir particles scattered throughout the axoplasm are first seen in the external layer of the ME on day 18.5 of gestation, and, on day 21.5 appear to terminate on the basement membrane of the perivascular space of the portal vessels. After birth, the fiber terminals contain several membrane-limited granules, which are labeled with ir PAP particles. Ultrathin, Epon-embedded sections of ME, treated by the protein A gold-labeling method for somatostatin, demonstrate positively labeled granules in the nerve fibers in the postnatal ME, but in the prenatal tissue, no specific gold-labeling is found. These findings show that, in the external layer of the ME, somatostatin storing occurs in the granules in the axonal terminals after birth.     

CCK-4-Induced Calcium Mobilization in T Cells Is Enhanced in Panic Disorder

Abstract: We investigated the effects of brain cholecystokinin (CCK) receptors on the intracellular calcium concentration and protein kinase C in human T cells. CCK-4 produced a transient increase in calcium in the absence of extracellular calcium. CCK-B agonists stimulated calcium mobilization in a dose-dependent manner in T cells. CCK-B antagonists suppressed CCK-4-induced calcium mobilization more potently than CCK-A antagonist. The recovery of desensitization of the CCK-4-induced response was delayed by a phosphoserine/phosphothreonine phosphatase inhibitor, calyculin A. The responsiveness to CCK-4 was also reduced by phorbol 12,13-dibutyrate (PDBu), and this effect of PDBu was abolished completely by preincubation with staurosporine. CCK-4-induced calcium mobilization was too small to attribute the desensitization to the protein kinase C transduction pathway. T cells from patients with untreated panic disorder exhibited significantly higher cholecystokinin-4-induced calcium mobilization than those from healthy controls or patients with treated panic disorder. These results suggest that cholecystokinin-B receptor function in T cells of patients with panic disorder is enhanced. Cholecystokinin-4-induced calcium mobilization in T cells may be state dependent and useful as a biological marker of panic disorder.     

A novel ferredoxin-dependent glutamate synthase from the hydrogen-oxidizing chemoautotrophic bacterium Hydrogenobacter thermophilus TK-6

Glutamate synthases are classified according to their specificities for electron donors. Ferredoxin-dependent glutamate synthases had been found only in plants and cyanobacteria, whereas many bacteria have NADPH-dependent glutamate synthases. In this study, Hydrogenobacter thermophilus, a hydrogen-oxidizing chemoautotrophic bacterium, was shown to possess a ferredoxin-dependent glutamate synthase like those of phototrophs. This is the first observation, to our knowledge, of a ferredoxin-dependent glutamate synthase in a nonphotosynthetic organism. The purified enzyme from H. thermophilus was shown to be a monomer of a 168-kDa polypeptide homologous to ferredoxin-dependent glutamate synthases from phototrophs. In contrast to known ferredoxin-dependent glutamate synthases, the H. thermophilus glutamate synthase exhibited glutaminase activity. Furthermore, this glutamate synthase did not react with a plant-type ferredoxin (Fd3 from this bacterium) containing a [2Fe-2S] cluster but did react with bacterial ferredoxins (Fd1 and Fd2 from this bacterium) containing [4Fe-4S] clusters. Interestingly, the H. thermophilus glutamate synthase was activated by some of the organic acids in the reductive tricarboxylic acid cycle, the central carbon metabolic pathway of this organism. This type of activation has not been reported for any other glutamate synthases, and this property may enable the control of nitrogen assimilation by carbon metabolism.     

Structure of thiocyanate hydrolase: a new nitrile hydratase family protein with a novel five-coordinate cobalt(III) center

Thiocyanate hydrolase (SCNase) of Thiobacillus thioparus THI115 is a cobalt(III)-containing enzyme catalyzing the degradation of thiocyanate to carbonyl sulfide and ammonia. We determined the crystal structures of the apo- and native SCNases at a resolution of 2.0 A. SCNases in both forms had a conserved hetero-dodecameric structure, (alphabetagamma)(4). Four alphabetagamma hetero-trimers were structurally equivalent. One alphabetagamma hetero-trimer was composed of the core domain and the betaN domain, which was located at the center of the molecule and linked the hetero-trimers with novel quaternary interfaces. In both the apo- and native SCNases, the core domain was structurally conserved between those of iron and cobalt-types of nitrile hydratase (NHase). Native SCNase possessed the post-translationally modified cysteine ligands, gammaCys131-SO(2)H and gammaCys133-SOH like NHases. However, the low-spin cobalt(III) was found to be in the distorted square-pyramidal geometry, which had not been reported before in any protein. The size as well as the electrostatic properties of the substrate-binding pocket was totally different from NHases with respect to the charge distribution and the substrate accessibility, which rationally explains the differences in the substrate preference between SCNase and NHase.     

Effects of a gonadotropin-releasing hormone antagonist on gonadotropin levels in Masu salmon and Sockeye salmon

The salmon gonadotropin-releasing hormone (sGnRH) is considered to be involved in gonadal maturation via gonadotropin (GTH) secretion in salmonid fishes. However, there is no direct evidence for endogenous sGnRH-stimulated GTH secretion in salmonids. In this study, to clarify whether endogenous sGnRH stimulates GTH secretion, we examined the effects of the mammalian GnRH (mGnRH) antagonist [Ac-Delta(3)-Pro(1), 4FD-Phe(2), D-Trp(3,6)]-mGnRH on luteinizing hormone (LH) levels in 0-year-old masu salmon Oncorhynchus masou and sockeye salmon Oncorhynchus nerka. First, the effects of the GnRH antagonist on LH release were examined in 0-year-old precocious male masu salmon. GnRH antagonist treatment for 3 hr significantly inhibited an increase in plasma LH levels that was artificially induced by exogenous sGnRH administration, indicating that the GnRH antagonist is effective in inhibiting LH release from the pituitary. Subsequently, we examined the effect of the GnRH antagonist on LH synthesis in 0-year-old immature sockeye salmon that were pretreated with exogenous testosterone for 42 days to increase the pituitary LH contents; the testosterone treatment did not affect the plasma LH levels. GnRH antagonist treatment slightly but significantly inhibited an increase in the testosterone-stimulated pituitary LH content levels. However, no significant differences in the plasma LH levels were observed between the GnRH antagonist-treated and control groups. These results suggest that endogenous sGnRH is involved in LH secretion in salmonid fishes.     

An ArsRC fusion protein enhances arsenate sensing and detoxification

Arsenical resistance (ars) operons encode genes for arsenic resistance and biotransformation. The majority are composed of individual genes, but fusion of ars genes is not uncommon, although it is not clear if the fused gene products are functional. Here we report identification of a four-gene ars operon from Paracoccus sp. SY that has two arsR-arsC gene fusions. ArsRC1 and ArsRC2 are related proteins that consist of an N-terminal ArsR arsenite (As(III))-responsive repressor with a C-terminal ArsC arsenate reductase. The other two genes in the operon are gapdh and arsJ. GAPDH, glyceraldehyde 3-phosphate dehydrogenase, forms 1-arseno-3-phosphoglycerate (1As3PGA) from 3-phosphoglyceraldehyde and arsenate (As(V)), ArsJ is an efflux permease for 1As3PGA that dissociates into extracellular As(V) and 3-phosphoglycerate. The net effect is As(V) extrusion and resistance. ArsRs are usually selective for As(III) and do not respond to As(V). However, the substrates and products of this operon are pentavalent, which would not be inducers of the operon. We propose that ArsRC fusions overcome this limitation by channelling the ArsC product into the ArsR binding site without diffusion through the cytosol, a de facto mechanism for As(V) induction. This novel mechanism for arsenate sensing can confer an evolutionary advantage for detoxification of inorganic arsenate.