High accumulation of elements in the human femoral artery

The relative contents (RCs) of elements in the femoral arteries as well as the thoracic aorta, coronary, basilar, and radial arteries from 26 subjects within the age range between 55 and 92 yr old, were analyzed by inductively coupled plasma atomic emission spectrometry. The RCs of calcium and phosphorus in the femoral arteries started to increase before the age of 60 yr. The RCs of magnesium increased after the age of 70 yr. However, the RCs of sulfur did not change significantly within the age range between 55 and 92 yr. With regard to localization of the mineral accumulations in the femoral arterial wall, it was found that the accumulations of calcium and phosphorus occurred only in the tunica media, only in the tunica intima, or in both the tunica media and the tunica intima. The manner of accumulation of calcium and phosphorus in the femoral arterial wall was different from that in the aortic wall. The average RCs of calcium in the 26 specimens were the highest in the femoral artery, followed in descending order by the thoracic aorta, coronary, basilar, and radial arteries. The average RCs of phosphorus were highest in the thoracic aorta, followed by the coronary, femoral, basilar, and radial arteries. It is noted that the accumulation of mineral elements never occurred uniformly in all the arteries.     

Dive Europa: a search-for-life initiative.

Liquid water, underwater volcanoes and possibly life forms have been suggested to be present beneath the estimated 10 km-thick ice shell of Europa the Jovian satellite J2. Europa's possible ocean is estimated to be 100-200km deep. Despite the great depth of the Europa's ocean, hydrostatic pressure at the seafloor would be 130-260 MPa, corresponding to 13-26 km depth of a theoretical Earth's ocean. The hydrostatic pressure is not beyond the edge of existing deep-sea technology. Here we propose exploration of Europa's deep-sea by the use of current technologies, taking a symbolic example of a deep submergence vehicle Shinkai 6500 which dives to a depth of 6.5 km deep (50 km depth of Europa's ocean). Shinkai 6500 is embarkable in the payload bay of the Space Shuttles in terms of size and weight for the transportation to a Low Earth Orbit (LEO). Secondary boost is needed for interplanetary flight from the LEO. On-orbit assembly of the secondary booster is a technological challenge. The International Space Station (ISS) and ISS-related technologies will facilitate the secondary boost. Also, ice shell drilling is a challenge and is needed before the dive into Europa's ocean. These challenges should be overcome during a certain leading time for matured experience in the ISS operation.     

Increased urinary catechol estrogen excretion in female smokers

Premenopausal female smokers show significantly increased estrogen 2-hydroxylation, which may account in part for the anti-estrogenic effects of cigarette smoking. We have measured five major urinary estrogens, including estradiol (E2), estrone (E1), 16 alpha-hydroxyestrone (16 alpha OHE1), estriol (E3), and 2-hydroxyestrone (2OHE1), in premenopausal female smokers and non-smokers, to determine whether increased C-2 hydroxylation affected the urinary excretory patterns in these subjects. While total measured estrogen excretion in the follicular phase did not differ significantly between the two groups, urinary 2OHE1 among the smokers constituted a significantly greater proportion of the total (31.1 vs 18.2%, P less than 0.02). This difference was largely caused by significantly increased urinary 2OHE1 and decreased E3 observed in smokers. A urinary catechol estrogen index, defined by [2OHE1]/[E3], was significantly elevated in smokers compared with non-smokers (1.67 +/- 0.21 vs 0.56 +/- 0.08, P less than 0.001), and this urinary index correlated strongly with radiometrically determined estrogen 2-hydroxylation (r = 0.84, P less than 0.01). Ratios of the various estrogen metabolites did not vary substantially throughout the menstrual cycle. Urinary estrogen indices as described here may therefore be useful in demonstrating differences in estrogen metabolism, specifically 2-hydroxylation vs 16 alpha-hydroxylation, in selected populations.     

Two Modes of Regulation of the Fatty Acid Elongase ELOVL6 by the 3-Ketoacyl-CoA Reductase KAR in the Fatty Acid Elongation Cycle

Fatty acids (FAs) are diverse molecules, and such diversity is important for lipids to exert their functions under several environmental conditions. FA elongation occurs at the endoplasmic reticulum and produces a variety of FA species; the FA elongation cycle consists of four distinct enzyme reactions. For this cycle to be driven efficiently, there must exist coordinated regulation of protein components of the FA elongation machinery. However, such regulation is poorly understood. In the present study, we performed biochemical analyses using the FA elongase ELOVL6 and the 3-ketoacyl-CoA reductase KAR, which catalyze the first and second steps of the FA elongation cycle, respectively. In vitro FA elongation assays using membrane fractions demonstrated that ELOVL6 activity was enhanced ∼10-fold in the presence of NADPH, although ELOVL6 itself did not require NADPH for its catalysis. On the other hand, KAR does use NADPH as a reductant in its enzyme reaction. Activity of purified ELOVL6 was enhanced by ∼3-fold in the presence of KAR. This effect was KAR enzyme activity-independent, since it was observed in the absence of NADPH and in the KAR mutant. However, ELOVL6 enzyme activity was further enhanced in a KAR enzyme activity-dependent manner. Therefore, KAR regulates ELOVL6 via two modes. In the first mode, KAR may induce conformational changes in ELOVL6 to become structure that can undergo catalysis. In the second mode, conversion of 3-ketoacyl-CoA to 3-hydroxyacyl-CoA by KAR may facilitate release of the product from the presumed ELOVL6–KAR complex.     

Relation between Neutral Lipid Accumulation and the Growth Phase in the Yeast,Lipomyces starkeyi,a Fat Producing Yeast

In order to investigate the substrate binding feature of undecaprenyl diphosphate synthase from Micrococcus luteus B-P 26 with respect to farnesyl diphosphate and a reaction intermediate, (Z,E,E)-geranylgeranyl diphosphate, we examined the reactivity of artificial substrate analogs, 3-desmethyl farnesyl diphosphate and 3-desmethyl Z-geranylgeranyl diphosphate, which lack the methyl group at the 3-position of farnesyl diphosphate and Z-geranylgeranyl diphosphate, respectively. Undecaprenyl diphosphate synthase did not accept either of the 3-desmethyl analogs as the allylic substrate, indicating that the methyl group at the 3-position of the allylic substrate is important in the undecaprenyl diphosphate synthase reaction. These analogs showed different inhibition patterns in the cis-prenyl chain elongation reaction with respect to the reactions of farnesyl diphosphate and Z-geranylgeranyl diphosphate as allylic substrate. These results suggest that the binding site for the natural substrate farnesyl diphosphate and those for the intermediate allylic diphosphate, which contains the cis-prenyl unit, are different during the cis-prenyl chain elongation reaction.     

Structural characterization of a rhamnose-binding glycoprotein (lectin) from Spanish mackerel (Scomberomorous niphonius) eggs

A rhamnose-binding glycoprotein (lectin), named SML, was isolated from the eggs of Spanish mackerel (Scomberomorous niphonius) by affinity and ion-exchange chromatographies. SML was composed of a non-covalently linked homodimer. The SML subunit was composed of 201 amino acid residues with two tandemly repeated domains, and contained 8 half-Cys residues in each domain, which is highly homologous to the N-terminal lectin domain of calcium-independent alpha-latrotoxin receptor in mammalian brains. Each domain has the same disulfide bonding pattern; Cys10-Cys40, Cys20-Cys99, Cys54-Cys86 and Cys67-Cys73 were located in the N-terminal domain, and Cys108-Cys138, Cys117-Cys195, Cys152-Cys182 and Cys163-Cys169 were in the C-terminal domain. SML was N-glycosylated at Asn168 in the C-terminal domain. The structure of the sugar chain was determined to be NeuAc-Galbeta1-4GlcNAcbeta1-2Manalpha1-6-(NeuAc-Galbeta1-4GlcNAcbeta1-2Manalpha1-3)Manbeta1-4GlcNAcbeta1-4GlcNAc-Asn.     

Inhibition of noradrenaline release from PC12 cells by the long-term treatment with cholera toxin

Guanine nucleotide-binding (G) proteins are required for intracellular vesicular transport and endocytosis. In this study, we investigated the effects of short-term (2 h) and long-term (24 h) treatment with cholera toxin (CTX), which ADP-ribosylates proteins having arginine residues such as the alpha subunit of Gs (G(s alpha)), on exocytosis from the neurosecretory rat pheochromocytoma PC 12 cell line. Short-term treatment with CTX stimulated the accumulation of cyclic AMP, and synergistically enhanced both extracellular Ca2+-dependent [3H]noradrenaline (NA) releases (induced by high K+ and ATP) and Ca2+-independent release (induced by mastoparan, a peptide in wasp venom). Long-term treatment with CTX for 24h inhibited Ca2+-dependent and -independent stimulated [3H]NA release. The inhibitory effect of long-term CTX treatment was not derived from a cyclic AMP-dependent system, because (1) H-89, an inhibitor of protein kinase A, had no effect on the inhibition induced by CTX, (2) the long-term treatment with forskolin did not show an inhibitory effect. [32P]ADP-ribosylation of G(s alpha) and its immunoreactivity with anti-G(s alpha) antiserum in the crude membrane fraction was inhibited in the long-term CTX-treated cells, but not in the long-term forskolin-treated cells. [32P]ADP-ribosylation of G(s alpha) in the membrane fraction of short-term CTX-treated cells was approximately 90% of the level in the control cells. These findings suggest that CTX stimulates [3H]NA release via a cyclic AMP-dependent system in the short-term, and that long-term CTX treatment inhibited its release, maybe via ADP-ribosylation of CTX-sensitive proteins such as G(s alpha).     

In vitro reconstitution of the GTPase-associated centre of the archaebacterial ribosome: the functional features observed in a hybrid form with Escherichia coli 50S subunits

We cloned the genes encoding the ribosomal proteins Ph (Pyrococcus horikoshii)-P0, Ph-L12 and Ph-L11, which constitute the GTPase-associated centre of the archaebacterium Pyrococcus horikoshii. These proteins are homologues of the eukaryotic P0, P1/P2 and eL12 proteins, and correspond to Escherichia coli L10, L7/L12 and L11 proteins respectively. The proteins and the truncation mutants of Ph-P0 were overexpressed in E. coli cells and used for in vitro assembly on to the conserved domain around position 1070 of 23S rRNA (E. coli numbering). Ph-L12 tightly associated as a homodimer and bound to the C-terminal half of Ph-P0. The Ph-P0.Ph-L12 complex and Ph-L11 bound to the 1070 rRNA fragments from the three biological kingdoms in the same manner as the equivalent proteins of eukaryotic and eubacterial ribosomes. The Ph-P0.Ph-L12 complex and Ph-L11 could replace L10.L7/L12 and L11 respectively, on the E. coli 50S subunit in vitro. The resultant hybrid ribosome was accessible for eukaryotic, as well as archaebacterial elongation factors, but not for prokaryotic elongation factors. The GTPase and polyphenylalanine-synthetic activity that is dependent on eukaryotic elongation factors was comparable with that of the hybrid ribosomes carrying the eukaryotic ribosomal proteins. The results suggest that the archaebacterial proteins, including the Ph-L12 homodimer, are functionally accessible to eukaryotic translation factors.     

Phylogenetic characterization of 16S rRNA gene clones from deep-groundwater microorganisms that pass through 0.2-micrometer-pore-size filters

A total of 247 clones of 16S rRNA genes from microorganisms captured by 0.2- and 0.1-microm-pore-size filters from sedimentary and granite rock aquifers were amplified and yielded 37 operational taxonomic units (OTUs). Fifteen OTUs captured by 0.1-microm-pore-size filters were affiliated with the candidate divisions OD1 and OP11, representing novel lineages. On the other hand, OTUs captured by 0.2-microm-pore-size filters were largely affiliated with Betaproteobacteria.     

Phylogenetic Characterization of 16S rRNA Gene Clones from Deep-Groundwater Microorganisms That Pass through 0.2-Micrometer-Pore-Size Filters

A total of 247 clones of 16S rRNA genes from microorganisms captured by 0.2- and 0.1-μm-pore-size filters from sedimentary and granite rock aquifers were amplified and yielded 37 operational taxonomic units (OTUs). Fifteen OTUs captured by 0.1-μm-pore-size filters were affiliated with the candidate divisions OD1 and OP11, representing novel lineages. On the other hand, OTUs captured by 0.2-μm-pore-size filters were largely affiliated with Betaproteobacteria.