Subcellular Localization of Enzymes of Carbon and Nitrogen Metabolism in Nodules of Medicago sativa

Alfalfa (Medicago sativa L. cv. Vernal) nodules were separatedinto host plant fractions and fractions of rhizobial originby differential centrifugation and sedimentation equilibriumcentrifugation. Both NAD- and NADP-linked isocitrate dehydrogenase(70%, 90%) and glutamate dehydrogenase activities (90%, 83%)were located primarily (percent total nodule activity) in thefractions of plant origin and their specific activities werehighest in the fractions of plant origin. More than 50% of thenodules' total activity of both glutamine synthetase and NAD-glutamatesynthase and greater than 90% of the total glutamate oxaloacetatetransaminase activity was located in plant fractions. However,the fractions of rhizobial origin had the highest specific activitiesof glutamine synthetase and glutamate synthase. (Received September 5, 1981; Accepted December 7, 1981)     

A New Assay for Determining Ganglioside Sialyltransferase Activities Lactosylceramide-2,3-Sialyltransferase (SAT I) and Monosialylganglioside-2,3-Sialyltransferase (SAT IV)

A new assay for the determination of lactosylceramide-2,3-sialyltransferase (SAT I, EC 2.4.99.9) and monosialoganglioside sialyltransferase (SAT IV, EC 2.4.99.2) is described. The assay utilised the commercially available fluorophore labelled sphingolipids, boron dipyrromethene difluoride (BODIPY) lactosylceramide (LacCer), and BODIPY-monosialotetrahexosylganglioside (GM1) as the acceptor substrates, for SAT I and SAT IV, respectively. HPLC coupled with fluorescence detection was used to analyse product formation. The analysis was performed in a quick and automated fashion. The assay showed good linearity for both BODIPY sphingolipids with a quantitative detection limit of 0.05 pmol. The high sensitivity enabled the detection of SAT I and SAT IV activities as low as 0.001 μU, at least 200 fold lower than that of most radiometric assays. This new assay was applied to the screening of SAT I and SAT IV activities in ovine and bovine organs (liver, heart, kidney, and spleen). The results provided evidence that young animals, such as calves, start to produce ganglioside sialyltransferases as early as 7 days after parturition and that levels change during maturation. Among the organs tested from a bovine source, spleen had the highest specific ganglioside sialyltransferase activity. Due to the organ size, the greatest total ganglioside sialyltransferase activities (SAT I and SAT IV) were detected in the liver of both bovine and ovine origin.     

Clinical adaptation of a high-performance liquid chromatographic method for the assay of pyridoxal 5′-phosphate in human plasma

Vitamin B6, measured as pyridoxal 5′-phosphate (PLP), is a co-enzyme in the transsulfuration pathway of homocysteine metabolism. Since depletion of PLP has been suggested as an independent risk factor for coronary artery disease, PLP is frequently measured to guide patient care. By a change and utilization of an Aquasil C18 column and the addition of an acetonitrile clean-up gradient to the potassium phosphate, with sodium perchlorate and bisulfite buffer between samples we report the modification of a previously described method for analysis of PLP. The result is a more practical, efficient, reliable and robust method for daily clinical use. We also determined and report that it is critical to protect freshly prepared standard PLP samples from light exposure during assay preparation.     

Cell-specific density of symbiotic dinoflagellates in tropical anthozoans

 Symbiotic dinoflagellates are abundant in the endoderm cells of tropical marine anthozoans, but the cell-specific density (CSD) of symbionts has not yet been investigated. In this study we used mechanical and enzymatic methods of maceration, and staining with substrate-specific fluorochromes, to observe a large number of individual host cells from 33 species of tropical anthozoans collected in Florida, Hawaii and Jamaica or cultured in Monaco. In the majority of species, most of the host cells contained a single algal cell (singlet). Host cells with two or more (up to six) algae were much less abundant. The average CSD for the 33 species was 1.54±0.30 (range 1.11 to 2.19). Singlets arranged in a monolayer can account for the areal density of algae observed in many anthozoans. The dinoflagellates occupy most of the interior of macerated host cells, leaving the host cytoplasm and cell membrane as a thin outer layer, often unresolvable by light microscopy. This spatial arrangement may favor diffusion and transport of CO₂, bicarbonate ions, and nutrients from the environment to the algae. The effect of nutrient enrichment on CSD was determined by exposing eleven species to chronically elevated levels of ammonium-N. After four weeks all species exhibited a dramatic increase in algal mitotic index and CSD. The potential consequences of environmentally induced increases in CSD in tropical anthozoans are discussed in terms of the decreased cell-specific photosynthesis (CO₂ limitation) and decreased rates of calcification observed in other studies. Accepted: 16 February 1998     

Bringing Home the Trash: Do Colony-Based Differences in Foraging Distribution Lead to Increased Plastic Ingestion in Laysan Albatrosses?

When searching for prey, animals should maximize energetic gain, while minimizing energy expenditure by altering their movements relative to prey availability. However, with increasing amounts of marine debris, what once may have been ‘optimal’ foraging strategies for top marine predators, are leading to sub-optimal diets comprised in large part of plastic. Indeed, the highly vagile Laysan albatross (Phoebastria immutabilis) which forages throughout the North Pacific, are well known for their tendency to ingest plastic. Here we examine whether Laysan albatrosses nesting on Kure Atoll and Oahu Island, 2,150 km apart, experience different levels of plastic ingestion. Twenty two geolocators were deployed on breeding adults for up to two years. Regurgitated boluses of undigestable material were also collected from chicks at each site to compare the amount of plastic vs. natural foods. Chicks from Kure Atoll were fed almost ten times the amount of plastic compared to chicks from Oahu despite boluses from both colonies having similar amounts of natural food. Tracking data indicated that adults from either colony did not have core overlapping distributions during the early half of the breeding period and that adults from Kure had a greater overlap with the putative range of the Western Garbage Patch corroborating our observation of higher plastic loads at this colony. At-sea distributions also varied throughout the year suggesting that Laysan albatrosses either adjusted their foraging behavior according to constraints on time away from the nest or to variation in resources. However, in the non-breeding season, distributional overlap was greater indicating that the energy required to reach the foraging grounds was less important than the total energy available. These results demonstrate how a marine predator that is not dispersal limited alters its foraging strategy throughout the reproductive cycle to maximize energetic gain and how this has led to differences in plastic ingestion.     

Alkaline hydrolysis and multiple site autophosphorylation differ for two forms of the epidermal growth factor receptor

Different tyrosines are autophosphorylated on the native and on the protease-generated 150 kDa forms of the epidermal growth factor receptor. High ATP concentrations increase the apparent molecular weight of already phosphorylated native receptors but not of the 150 kDa form, indicating that only the native receptor has multiple autophosphorylation sites available. The non-identity of the tyrosine-phosphates on the native and 150 kDa receptor forms is seen in their response to alkaline hydrolysis (10% and 40% resistant, respectively). Since the liberated phosphate is peptide bound, the native receptor fails to be alkali-resistant because of which peptide bonds are hydrolyzed.     

Integrating phenotype ontologies across multiple species

Phenotype ontologies are typically constructed to serve the needs of a particular community, such as annotation of genotype-phenotype associations in mouse or human. Here we demonstrate how these ontologies can be improved through assignment of logical definitions using a core ontology of phenotypic qualities and multiple additional ontologies from the Open Biological Ontologies library. We also show how these logical definitions can be used for data integration when combined with a unified multi-species anatomy ontology.