We describe a sheath flow capillary electrophoresis timeof-flight mass spectrometry (CE-TOFMS) method in the negative mode using a platinum electrospray ionization (ESI) spray needle, which allows the comprehensive analysis of anionic metabolites. The material of the spray needle had significant effect on the measurement of anions. A stainless steel spray needle was oxidized and corroded at the anodic electrode due to electrolysis. The precipitation of iron oxides (rust) plugged the capillary outlet, resulting in shortened capillary lifetime. Many anionic metabolites also formed complexes with the iron oxides or migrating nickel ion, which was also generated by electrolysis and moved toward the cathode (the capillary inlet). The metal-anion complex formation significantly reduced detection sensitivity of the anionic compounds. The use of a platinum ESI needle prevented both oxidation of the metals and needle corrosion. Sensitivity using the platinum needle increased from several-to 63-fold, with the largest improvements for anions exhibiting high metal chelating properties such as carboxylic acids, nucleotides, and coenzyme A compounds. The detection limits for most anions were between 0.03 and 0.87 µmol/L (0.8 and 24 fmol) at a signal-to-noise ratio of 3. This method is quantitative, sensitive, and robust, and its utility was demonstrated by the analysis of the metabolites in the central metabolic pathways extracted from mouse liver.Metabolism is the entire network of chemical reactions that occur in a cell in order to maintain life, in which one metabolite is transformed into another by a sequence of enzymes. Among the whole cellular metabolic network, central carbon metabolism, composed of glycolysis, the pentose phosphate pathway, and the tricarboxylic acid (TCA) cycle, plays key functions in substrate degradation, energy and cofactor regeneration, and biosynthetical precursor supply (DNA, RNA, proteins, peptideglycan, and lipid bilayers).1,2 Interestingly, all the components involved in the central carbon and energy metabolism are negatively charged: phosphorylated saccharides, phosphorylated carboxylic acids, carboxylic acids, coenzyme A (CoA) compounds, nucleotides, and nicotinamide adenine dinucleotides.As the importance of metabolomics is recognized, several largescale metabolite analysis methods using GC/MS, 3 LC/MS,

Studies of the halide complexation of element 105 in aqueous solution were performed on 34-s 262 Ha produced in the 249 Bk( ls O,5n) reaction. The 262 Ha was detected by measuring the fission and alpha activities associated with its decay and the alpha decays of its daughter, 4.3-s 258 Lr. Time-correlated pairs of parent and daughter alpha particles provided a unique identification of the presence of 262 Ha. About 1600 anion exchange separations of 262 Ha from HCl and mixed HC1/HF solutions were performed on a one-minute time scale. Reversed-phase micro-chromatographic columns incorporating triisooctyl amine (TIOA) on an inert support were used in the computer-controlled liquid chromatography apparatus, ARCA II. -.

We report the identification and characterization of a novel 124-kDa Ran binding protein, RanBP5. This protein is related to importin-␤, the key mediator of nuclear localization signal (NLS)-dependent nuclear transport. RanBP5 was identified by two independent methods: it was isolated from HeLa cells by using its interaction with RanGTP in an overlay assay to monitor enrichment, and it was also found by the yeast two-hybrid selection method with RanBP1 as bait. RanBP5 binds to RanBP1 as part of a trimeric RanBP1-Ran-RanBP5 complex. Like importin-␤, RanBP5 strongly binds the GTP-bound form of Ran, stabilizing it against both intrinsic and RanGAP1-induced GTP hydrolysis and also against nucleotide exchange. The GAP resistance of the RanBP5-RanGTP complex can be relieved by RanBP1, which might reflect an in vivo role for RanBP1. RanBP5 is a predominantly cytoplasmic protein that can bind to nuclear pore complexes. We propose that RanBP5 is a mediator of a nucleocytoplasmic transport pathway that is distinct from the importin-␣-dependent import of proteins with a classical NLS.Ran/TC4 is a highly abundant, small GTP-binding and -hydrolyzing protein that is located predominantly in the nucleus (8,12). Ran has the biochemical properties of a GTPase switch cycling between two conformational states, the GTP-bound state and the GDP-bound state. The intrinsic rates of nucleotide exchange and GTP hydrolysis are very low and can be increased up to five orders of magnitude by the regulatory proteins RCC1 and RanGAP1, respectively (4,8,22). The first indications concerning the functions of Ran came from the analysis of mutants of these Ran regulators in both mammalian and yeast cells. These studies implicated Ran in a variety of processes, including the onset of mitosis, initiation of S phase, exit from mitosis, maintenance of nuclear structure, and premRNA processing and mRNA export into the cytoplasm (for reviews, see references 41 and 43).Furthermore, in vitro studies with permeabilized cells showed that Ran is an essential factor for the nuclear localization signal (NLS)-dependent nuclear protein import (30, 31). Macromolecular transport across the nuclear envelope occurs at the nuclear pore complexes (NPCs) and involves the import of proteins into the cell nucleus and the export of RNAs and proteins. Four soluble cytosolic factors are required to reconstitute nuclear import of NLS substrates in vitro. In addition to Ran, these are importin-␣ and importin-␤ (also known as karyopherin-␣ and karyopherin-␤) (17,19,36) and NTF2 (alternatively known as pp15 or p10) (32, 34). Together, importin-␣ and importin-␤ comprise the import receptor complex, where importin-␣ binds proteins bearing an NLS and importin-␤ mediates the interaction with the NPC. Ran appears to be required for at least two steps in nuclear import. First, translocation through the nuclear pore requires GTP hydrolysis by Ran, probably even as the sole source of energy (47). Second, the disassembly of the importin-␣/-␤-NLS protein complex following translocation ...