Large Inversion in Escherichia Coli K-12 1485in between Inversely Oriented Is3 Elements near Lac and Cdd

A companion study has shown that the inversion carried by strain 1485IN has one terminus between lac and proC and the other between his and cdd of the normal strain. Starting with this mapping data, we have done molecular work demonstrating that the inversion occurred by recombination between inversely oriented two IS3 elements, one present near lac and the other near the cdd locus; i.e., the inversion is IN(is3B-is3E). Evidence supporting this conclusion includes: (i) Normal and inversion strains share two short regions with identical restriction maps. One of these regions is near lac and the other near cdd. (ii) IS3 homology was detected in each of the terminus regions of both the normal and inversion strains. (iii) The sequence on one side of the original IS3 element near lac has been exchanged with the sequence on one side of the IS3 near cdd. Whether the inversion has occurred by one event of homologous recombination between the two IS3 elements or has been caused by involvement of IS3 elements on an F factor is discussed. Another rearrangement, probably related to inversion and deletion, was detected between the IS3 and cdd of the inversion strain.     

Site-specific recombinase genes in three Shigella subgroups and nucleotide sequences of a pinB gene and an invertible B segment from Shigella boydii.

Inversional switching systems in procaryotes are composed of an invertible DNA segment and a site-specific recombinase gene adjacent to or contained in the segment. Four related but functionally distinct systems have previously been characterized in detail: the Salmonella typhimurium H segment-hin gene (H-hin), phage Mu G-gin, phage P1 C-cin, and Escherichia coli e14 P-pin. In this article we report the isolation and characterization of three new recombinase genes: pinB, pinD, and defective pinF from Shigella boydii, Shigella dysenteriae, and Shigella flexneri, respectively. The genes pinB and pinD were detected by the complementation of a hin mutation of Salmonella and were able to mediate inversion of the H, P, and C segments. pinB mediated H inversion as efficiently as the hin gene did and mediated C inversion with a frequency three orders of magnitude lower than that of the cin gene. pinD mediated inversion of H and P segments with frequencies ten times as high as those for the genes intrinsic to each segment and mediated C inversion with a frequency ten times lower than that for cin. Therefore, the pinB and pinD genes were inferred to be different from each other. The invertible B segment-pinB gene cloned from S. boydii is highly homologous to the G-gin in size, organization, and nucleotide sequence of open reading frames, but the 5' constant region outside the segment is quite different in size and predicted amino acid sequence. The B segment underwent inversion in the presence of hin, pin, or cin. The defective pinF gene is suggested to hae the same origin as P-pin on e14 by the restriction map of the fragment cloned from a Pin+ transductant that was obtained in transduction from S. flexneri to E. coli delta pin.     

Biosynthesis of poly(3-hydroxyalkanoate) from amino acids

It was found that an optically active copolyester, poly(3-hydroxybutyrate-co-3-hydroxyvalerate), denoted as P(3HB-co-3HV), is synthesized by Alcaligenes eutrophus H16 from several amino acids under various fermentation conditions. The optimum condition for the biosynthesis from one amino acid, threonine, was investigated and its biosynthetic pathway was discussed on the basis of the relation between the fermentation condition and the co-monomer composition of the produced polyesters.     

Enhancement of DNA synthesis in rat thymocytes by stimulating their muscarinic acetylcholine receptors.

The binding of 3H-acetylcholine (ACh) to acetylcholine receptors (AChRs) on rat thymocytes was examined and found to be inhibited by the treatment with several antagonists against nicotinic and muscarinic AChRs. This result suggested that thymocytes have AChRs with different affinity, and bear both nicotinic and muscarinic AChRs on their surfaces. To make clear the functional significance of the AChRs, DNA synthesis of the thymocytes stimulated with ACh was examined. 3H-thymidine uptake of thymocytes was significantly increased when the cells were stimulated with ACh or another cholinergic agonist. The increment of DNA synthesis caused by ACh in thymocytes was not reduced by treatment with nicotinic antagonists, but was decreased by treatment with any of the muscarinic antagonists. Concentration of the intracellular second messengers, inositol 1,4,5-triphosphate (IP3) and guanosine 3',5'-cyclic monophosphate (cGMP) was also made higher by ACh stimulation. It is discussed that the enhancement of intracellular IP3 and cGMP concentrations after stimulation of muscarinic AChRs appears to be related with the increment of thymocyte DNA synthesis.     

Structural differences among guinea pig Fc gamma 1/gamma 2 receptors on macrophages, polymorphonuclear cells, and lymphocytes.

Using the cDNA, D-3, coding for Fc gamma 1/gamma 2 receptor of guinea pig macrophages that binds IgG1 and IgG2 (Fc gamma 1/gamma 2R), we examined the cell distribution of this receptor by RNA blot analysis. The Fc gamma 1/gamma 2R mRNA was expressed in polymorphonuclear cells and B cells as well as in macrophages, but not at the detectable level in T cells. The cDNA amplified from RNA of polymorphonuclear cells in the polymerase chain reaction was the same as D-3. The cDNA of B cells was found to have about 140 bp cDNA segment inserted to the cytoplasmic tail of D-3. We found that the cDNA amplified from T cell RNA differed in signal peptide and extracellular domain sequence from cDNAs of other cell types. This cDNA does not seem to be amplified from the mRNAs of contaminating other cell types.     

Application of immobilized lipase to regio-specific interesterification of triglyceride in organic solvent

Summary Lipase from Rhizopus delemar was immobilized by entrapment with photo-crosslinkable resin prepolymers or urethane prepolymers or by binding to various types of porous silica beads. The immobilized lipase preparations thus obtained were examined for their activity in converting olive oil to an interesterified fat (cacao butter-like fat), whose oleic acid moieties at 1- and 3-positions were replaced with stearic acid moieties, in the reaction solvent n-hexane. Although all of the immobilized preparations exhibited some activity, lipase adsorbed on Celite and then entrapped with a hydrophobic photo-crosslinkable resin prepolymer showed the highest activity, about 75% of that of lipase simply adsorbed onto Celite. Entrapment markedly enhanced the operational stability of lipase.Dedicated to Professor H. Holzer, Freiburg University, on his 60th birthday (June 13, 1981)     

Seasonal and areal features of the lagoonal environment in Lake Nakanoumi,a shallow coastal lagoon in Japan

Lake Nakanoumi is a shallow coastal lagoon connected with the Japan Sea by a narrow channel. Over the past decade, land reclamation resulted in a 33% reduction of the lagoon's surface area. The remaining water basin of Lake Nakanoumi is scheduled to be artificially freshened to supply irrigation water for the newly reclaimed lands. This paper deals with the seasonal and areal features of the lagoonal environment prior to the beginning of the artificial desalinization.     

Simultaneous determination of triazolo-benzophenone [2′,5-dichloro-2-(3-glycylaminomethyl-5-methyl-4H-1,2,4-triazol-4-yl)-benzophenone] and its major blood metabolite, triazolam, in monkey plasma by electron-capture gas—liquid chromatography

A gas—liquid chromatographic method for the simultaneous determination of triazolobenzophenone [2′,5-dichloro-2-(3-glycylaminomethyl-5-methyl-4H-1,2,4-triazol-4-yl)-benzophenone, TB] and its major blood metabolite, triazolam, 8-chloro-6-(o-chlorophenyl)-1-methyl-4H-s-triazolo[4,3-a][1,4]benzodiazepine (TZ), in monkey plasma was developed. Decomposition of TB was observed during gas—liquid chromatography. In alkaline medium, TB in plasma was submitted to ring closure reaction to yield triazolo-aminoquinoline, [4-amino-7-chloro-5-(2-chlorophenyl)-1-methyl-4H-s-triazolo[4,3-a]quinoline (TAQ), while TZ remained unaffected, and TAQ and TZ in the benzene extract were assayed by gas—liquid chromatography using an electron-capture detector. The concentration ranges studied were from 5 to 40 ng of TB per 0.5 ml of plasma and from 2 to 20 ng of TZ per 0.5 ml of plasma. This method could be applied to the determination of the plasma levels of TB and TZ in monkeys following intravenous administration of a single 0.2 mg/kg dose of TB.     

Indole-3-acetaldehyde oxidase of pea seedlings

Indole-3-acetaldehyde oxidase was partially purified from the epicotyl of Pisum satiyum seedlings by column chromatography using CM-Sephadex and Sephadex G-150. The enzyme was only active in the presence of molecular oxygen. The activity was maximal at pH 8.0, and the Km value for indole-3-acetaldehyde was 1.4 × 10⁻³ M . The enzyme was inhibited strongly by p -hydroxymercuribenzoate, cyanide and hydroxylamine, suggesting that it contains sulfhydryl group(s) and a metal component such as iron.     

Auxin-induced changes in the cell wall structure: Changes in the sugar compositions, intrinsic viscosity and molecular weight distributions of matrix polysaccharides of the epicotyl cell wall of Vigna angularis

Rapid effects of indole-3-acetic acid (IAA) on the mechanical properties of cell wall, and sugar compositions, intrinsic viscosity and molecular weight distribution of cell wall polysaccharides were investigated with excised epicotyl segments of Vigna angularis Ohwi et Ohashi cv. Takara.

• 1 IAA caused cell wall loosening as studied by stress-relaxation analysis within 15 min after the IAA application.
• 2 IAA stimulated the decrease in the content of arabinose and galactose in the hemicellulose 1 h after its application. The amounts of other component sugars in the cell wall polysaccharides remained constant during the IAA-induced segment growth.
• 3 The intrinsic viscocity of the pectin increased as early as 30 min after the IAA application. This effect was not prevented when elongation growth of the segment was osmotically suppressed by 0.15 M mannitol.
• 4 Gel permeation chromatography of the pectin on a Sepharose 4 B column demonstrated that IAA caused increase in the mass-average molecular weight of the pectin. Analysis of the sugar compositions of the pectin eluted from the Sepharose 4 B column indicated that IAA increased the molecular weight of the polysaccharides composed of uronic acid, galactose, rhamnose and arabinose. This effect became apparent within 30 min after the IAA application. Furthermore, IAA increased the molecular weight of the pectin when elongation growth of the epicotyl segments was osmotically suppressed by 0.15 M mannitol.
• 5 Hemicellulose of the cell wall chromatographed on a Sepharose CL-4 B column. Analysis of the neutral sugar compositions and the iodine staining property (specific for xyloglucans) of the polysaccharide solution eluted from the column indicated that the hemicellulose consisted of xyloglucans, arabinogalactans and polysaccharides composed of xylose and/or mannose. IAA caused a decrease in the arabinogalactan content and depolymerization of xyloglucans. These IAA effects became apparent within 30 min after the IAA application. These changes occurred even when elongation growth of the epicotyl segments was osmotically suppressed by 0.15 M mannitol.

Polymerization of the pectin, degradation of arabinogalactans and depolymerization of xyloglucans appear to be involved in the mechanism by which IAA induces cell wall loosening and therefore extension growth of cells.