High-performance anion-exchange chromatography of ultraviolet-absorbing constituents of human urine

A 100-μl urine sample was chromatographed on a column packed with a strongly basic macroreticular anion-exchange resin (Diaion CDR-10, 5– μm diameter with a nominal 35% cross linkage). The elution was performed with a linear acetate gradient from 0 to 6.0 M at an average flow-rate of 0.72 ml/min and at an average pressure of 104 kg/cm². The relative standard deviation of retention times and peak height was ± 4% or less. The properties of the macroreticular anion-exchange resin, the effect of the particle size, the pH of acetate buffers, and the effect of the flow-rate of the eluent on the separation were investigated. Thirty three components of urine were then resolved and named.     

Hydrogen evolution by co-immobilized Chlorella vulgaris and Clostridium butyricum cells

Whole cells of Chlorella vulgaris and Clostridium butyricum were co-immobilized in 2% agar gel. NADP was suitable as an electron carrier. The rate of hydrogen evolution increased with increasing NADP concentration. The optimum conditions for hydrogen evolution were pH 7.0 and 37°C. The immobilized C. vulgaris-NADP-immobilized Cl. butyricum system continuously evolved hydrogen at a rate of 0.29–1.34 μmol/h per mg Chl for 6 days. On the other hand, the system without NADP evolved only a trace amount of hydrogen.     

Sequence of an intestinal cDNA encoding human motilin precursor

A cDNA clone encoding the human motilin precursor was isolated from an intestinal library using synthetic oligonucleotide probes. The predicted amino acid sequence indicates that the motilin precursor consists of 115 amino acids and includes a 25-residue N-terminal signal peptide followed by the 22-amino-acid motilin sequence and a long, 68-residue C-terminal peptide. The amino acid sequence of human motilin predicted from the cDNA sequence is identical to its porcine counterpart, which has been determined by protein sequencing. Proteolytic processing of promotilin to motilin occurs at the sequence, Lys-Lys, this being the first reported instance of processing occurring at a pair of Lys residues. In other precursors it occurs at Lys-Arg, Arg-Arg, Arg, or very rarely Lys.     

Production of a herbicide, 5-aminolevulinic acid,by Rhodobacter sphaeroides using the effluent of swine waste from an anaerobic digestor

Summary For the production of a herbicide, 5-amino-levulinic acid (ALA), from anaerobic digestion liquor, the utilization of the photosynthetic bacterium, Rhodobacter sphaeroides was examined. This bacterium could produce ALA extracelularly from this liquor with the addition of levulinic acid (LA), an inhibitor of ALA dehydratase (ALAD), and glycine, a precursor of ALA biosynthesis in the Shemin pathway. Succinate (another precursor) addition was unnecessary for ALA production. When repeated additions of LA were made together with glycine ALA production was significantly enhanced. However, above three additions of LA, ALA production was not further enhanced. The maximum value of ALA production attained was 4.2 mM (0.63 g/ 1), which was over double that of other ALA producers such as Chlorella vulgaris. Propionic acid was predominantly utilized compared with other lower fatty acids, suggesting that this might be converted to ALA via succinyl-coenzyme A (CoA) in the methylmalonyl-CoA pathway.Offprint requests to: Y. Nishizawa     

A photochemical fuel cell system using Anabaena N-7363

Summary A blue-green algae, Anabaena N-7363, was immobilized in 2% agar gel. The hydrogen productivity of the immobilized algae was three times higher than that of free algae. The maximum hydrogen production rate by the immobilized blue-green algae was 0.52 moles h^–1 g^–1 (of wet gel) in the medium without nitrogen sources under illumination (10,000 lux). The oxygen evolved was then removed by a reactor containing aerobic bacteria. A photo-current of 15–20 mA was continuously produced for 7 days by the photochemical fuel cell system consisting of the immobilized Anabaena reactor, the oxygen-removing reactor and the hydrogen-oxygen fuel cell. The conversion ratio of hydrogen to current was from 80% to 100%.     

Nucleotide sequence of the genes for β and ε subunits of proton-translocating ATPase from Escherichia coli

The 1855-nucleotide long DNA sequence of part of the gene cluster for the proton-translocating ATPase from $\text{E. coli}$ was determined by the method of Maxam-Gilbert. The sequence covers the genes for the β and ε subunits of F₁ along with the flanking region. The amino acid sequence of these subunits deduced from the nucleotide sequence indicates that the β and ε subunits have 459 and 138 amino acids, respectively. The possible secondary structure of the both subunits was estimated from the deduced primary structures. A possible nucleotide binding site in the β subunit is also discussed on the basis of the primary and secondary structures. The codons used in the genes for all the components of F₁F₀ were different in different genes, suggesting that the amount of each subunit in the F₁F₀ is determined to some extent on a translational level.     

Determination of eperisone in human plasma by gas chromatography—mass spectrometry

A gas chromatographic—mass spectrometric method was developed to determine eperisone hydrochloride, 4′-ethyl-2-methyl-3-piperidinopropiophenone hydrochloride, in human plasma over the concentration range 0.2–40 ng/ml. Excellent sensitivity was achieved by selection of a favorable fragment ion, m/z 98, of eperisone and reduction of heat decomposition of eperisone by using a splitless injector and a shortened capillary column. The method described here allows the determination of plasma concentrations as low as 0.2 ng/ml, the concentration attained 6 h after a single oral administration of 50 mg. At eperisone hydrochloride concentrations higher than 0.5 ng/ml, the mean inter-day variation of accuracy of the assay was less than 12%.     

NDR2‐mediated Rabin8 phosphorylation is crucial for ciliogenesis by switching binding specificity from phosphatidylserine to Sec15

Primary cilia are antenna‐like sensory organelles protruding from the plasma membrane. Defects in ciliogenesis cause diverse genetic disorders. NDR2 was identified as the causal gene for a canine ciliopathy, early retinal degeneration, but its role in ciliogenesis remains unknown. Ciliary membranes are generated by transport and fusion of Golgi‐derived vesicles to the pericentrosome, a process requiring Rab11‐mediated recruitment of Rabin8, a GDP–GTP exchange factor (GEF) for Rab8, and subsequent Rab8 activation and Rabin8 binding to Sec15, a component of the exocyst that mediates vesicle tethering. This study shows that NDR2 phosphorylates Rabin8 at Ser‐272 and defects in this phosphorylation impair preciliary membrane assembly and ciliogenesis, resulting in accumulation of Rabin8‐/Rab11‐containing vesicles at the pericentrosome. Rabin8 binds to and colocalizes with GTP‐bound Rab11 and phosphatidylserine (PS) on pericentrosomal vesicles. The phospho‐mimetic S272E mutation of Rabin8 decreases affinity for PS but increases affinity for Sec15. These results suggest that NDR2‐mediated Rabin8 phosphorylation is crucial for ciliogenesis by triggering the switch in binding specificity of Rabin8 from PS to Sec15, thereby promoting local activation of Rab8 and ciliary membrane formation.     

Research Commentary: Association of Zoonotic Pathogens with Fresh, Estuarine, and Marine Macroaggregates

Aquatic macroaggregates (flocs ≥0.5 mm) provide an important mechanism for vertical flux of nutrients and organic matter in aquatic ecosystems, yet their role in the transport and fate of zoonotic pathogens is largely unknown. Terrestrial pathogens that enter coastal waters through contaminated freshwater runoff may be especially prone to flocculation due to fluid dynamics and electrochemical changes that occur where fresh and marine waters mix. In this study, laboratory experiments were conducted to evaluate whether zoonotic pathogens (Cryptosporidium, Giardia, Salmonella) and a virus surrogate (PP7) are associated with aquatic macroaggregates and whether pathogen aggregation is enhanced in saline waters. Targeted microorganisms showed increased association with macroaggregates in estuarine and marine waters, as compared with an ultrapure water control and natural freshwater. Enrichment factor estimations demonstrated that pathogens are 2–4 orders of magnitude more concentrated in aggregates than in the estuarine and marine water surrounding the aggregates. Pathogen incorporation into aquatic macroaggregates may influence their transmission to susceptible hosts through settling and subsequent accumulation in zones where aggregation is greatest, as well as via enhanced uptake by invertebrates that serve as prey for marine animals or as seafood for humans.