生活援助方法論の教育方法とその考え方

教育活動Educational Activity授業科目「生活援助方法論I・II」の教育目標・教育内容および科目構造、授業の展開にあたって基盤としている考え方、生活援助方法論Iの具体的な授業計画を紹介した。また、最初の授業の導入方法はその後の学修に大きな影響を及ぼすといわれるが、入学直後の学生に実施した授業の導入と、学生の気づきや学びについて報告する

Indium-Catalyzed Annulation of o-Acylanilines with Alkoxyheteroarenes: Synthesis of Heteroaryl[b]quinolines and Subsequent Transformation to Cryptolepine Derivatives

We disclose herein the first synthetic method that is capable of offering heteroaryl[b]quinolines (HA[b]Qs) with structural diversity, which include tricyclic and tetracyclic structures with (benzo)thienyl, (benzo)furanyl, and indolyl rings. The target HA[b]Q is addressed by the annulation of o-acylanilines and MeO–heteroarenes with the aid of an indium Lewis acid that effectively works to make two different types of the N–C and C–C bonds in one batch. A series of indolo[3,2-b]quinolines prepared here can be subsequently transformed to structurally unprecedented cryptolepine derivatives. Mechanistic studies showed that the N–C bond formation is followed by the C–C bond formation. The indium-catalyzed annulation reaction thus starts with the nucleophilic attack of the NH2 group of o-acylanilines to the MeO-connected carbon atom of the heteroaryl ring in an SNAr fashion, and thereby the N–C bond is formed. The resulting intermediate then cyclizes to make the C–C bond through the nucleophilic attack of the heteroaryl-ring-based carbon atom to the carbonyl carbon atom, providing the HA[b]Q after aromatizing dehydration

Crystal structure of the Hfq and catalase HPII complex.

<p>(A) Stereo diagram showing the crystal packing of the complex composed of Hfq hexamers in cyan and HPII tetramers in violet. All the four bound Hfq hexamers are displayed for the HPII tetramer at the center, whereas only two hexamers are displayed for each surrounding HPII tetramer for clarity. (B) Structure of one HPII tetramer with four bound Hfq hexamers showing interaction through their distal surfaces. Viewed in stereo. Subunits of one Hfq hexamer are displayed in cyan and green. Numbers 2 – 5 indicate the subunit number in the Hfq hexamer as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078216#pone-0078216-g004" target="_blank">Fig. 4</a>. One molecule of the HPII tetramer is displayed in tan (the C-terminal lobe) and in magenta (the other parts). A space-filling model in blue represents heme. Other models are in grey.</p

Post-Transcriptional Regulator Hfq Binds Catalase HPII: Crystal Structure of the Complex

<div><p>We report a crystal structure of Hfq and catalase HPII from <i>Escherichia coli</i>. The post-transcriptional regulator Hfq plays a key role in the survival of bacteria under stress. A small non-coding RNA (sRNA) DsrA is required for translation of the stationary phase sigma factor RpoS, which is the central regulator of the general stress response. Hfq facilitates efficient translation of <i>rpoS</i> mRNA, which encodes RpoS. Hfq helps in the function of other specific proteins involved in RNA processing, indicating its versatility in the cell. However, structural information regarding its interactions with partners is missing. Here we obtained crystals of Hfq and HPII complexes from cell lysates following attempts to overexpress a foreign membrane protein. HPII is one of two catalases in <i>E. coli</i> and its mRNA is transcribed by an RNA polymerase holoenzyme containing RpoS, which in turn is under positive control of small non-coding RNAs and of the RNA chaperone Hfq. This sigma factor is known to have a pronounced effect on the expression of HPII. The crystal structure reveals that a Hfq hexamer binds each subunit of a HPII tetramer. Each subunit of the Hfq hexamer exhibits a unique binding mode with HPII. The hexamer of Hfq interacts via its distal surface. The proximal and distal surfaces are known to specifically bind different sRNAs, and binding of HPII could affect Hfq function. Hfq-HPII complexation has no effect on catalase HPII activity.</p></div

Interactions between Hfq subunit 5 and HPII.

<p>Subunit 5 is displayed in cyan and one molecule of the HPII tetramer is color-coded as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078216#pone-0078216-g004" target="_blank">Fig. 4A</a>. Other models are displayed in grey. Interacting residues are drawn in ball-and-stick representation overlaid with 2 Fo - Fc maps of 1.0 σ, and atoms are color-coded as: nitrogen, blue; oxygen, red; and carbon, yellow in Hfq and magenta in HPII. Bonds are depicted as black dashes. Interactions of subunits 2 and 6 with HPII are shown in Fig. S4 in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078216#pone.0078216.s001" target="_blank">File S1</a> and those of subunits 1 and 4 are in Fig. S5 in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078216#pone.0078216.s001" target="_blank">File S1</a>. See also Table S2 in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078216#pone.0078216.s001" target="_blank">File S1</a> for bond length and type.</p

Characterization of protein samples.

<p>(A) An SDS-PAGE pattern of isolated protein solution and crystals of the Hfq and catalase HPII complex dissolved in SDS buffer solution. Lanes are: 1, markers; 2, protein solution after nickel-affinity chromatography; and 3, crystals of Hfq and HPII complex. Arrows and numbers in lane 3 indicate samples for PMF MALDI-TOF analysis. Asterisk in lane 2 indicates an overproduced PomA protein with a histidine tag. Bands 1 and 2 were identified as dimer and monomer of HPII and bands 3 and 4 as hexamer and monomer of Hfq by the PMF analysis, respectively. Note that the fraction eluting from the nickel-affinity column contained significant amounts of both Hfq and HPII. (B) Typical electron micrographs of complexes of endogenous Hfq and HPII prepared with negative staining. Some ring-like structures with dimensions of 70 – 80 Å are indicated with arrows. Bar indicates 100 Å.</p

Some structural details.

<p>(A) Two HPII molecules and their interaction partners, two Hfq hexamers. Hfq subunits are color-coded as in Figs. 3B and 4B, and HPII is displayed in tan (the C-terminal lobe) and in violet (the other parts) with heme in blue. “P” and “D” denote the proximal and distal sides of the Hfq hexamer, respectively. (B) Hfq hexamer viewed from the distal side. Residues for binding to HPII are drawn in space-filling representation (see also Table S2 in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078216#pone.0078216.s001" target="_blank">File S1</a>) with the single-letter amino acid code for Tyr 25 and Asn 28 in subunit 5 (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0078216#pone-0078216-g005" target="_blank">Fig. 5</a>). Atoms are color-coded as: carbon, yellow; nitrogen, blue; and oxygen, red. “α1” denotes the N-terminal α-helix and “β1” – “β5” β-strands. Numbers 1 - 6 in A and B indicate the subunit number in the Hfq hexamer. Only the Hfq subunits on the front side have the number in A for clarity.</p

Biochemical and Molecular Characterization of a Novel UDP-Glucose:Anthocyanin 3′-O-Glucosyltransferase, a Key Enzyme for Blue Anthocyanin Biosynthesis, from Gentian

Gentian (Gentiana triflora) blue petals predominantly contain an unusually blue and stable anthocyanin, delphinidin 3-O-glucosyl-5-O-(6-O-caffeoyl-glucosyl)-3′-O-(6-O-caffeoyl-glucoside) (gentiodelphin). Glucosylation and the subsequent acylation of the 3′-hydroxy group of the B-ring of anthocyanins are important to the stabilization of and the imparting of bluer color to these anthocyanins. The enzymes and their genes involved in these modifications of the B-ring, however, have not been characterized, purified, or isolated to date. In this study, we purified a UDP-glucose (Glc):anthocyanin 3′-O-glucosyltransferase (3′GT) enzyme to homogeneity from gentian blue petals and isolated a cDNA encoding a 3′GT based on the internal amino acid sequences of the purified 3′GT. The deduced amino acid sequence indicates that 3′GT belongs to the same subfamily as a flavonoid 7-O-glucosyltransferase from Schutellaria baicalensis in the plant glucosyltransferase superfamily. Characterization of the enzymatic properties using the recombinant 3′GT protein revealed that, in contrast to most of flavonoid glucosyltransferases, it has strict substrate specificity: 3′GT specifically glucosylates the 3′-hydroxy group of delphinidin-type anthocyanins containing Glc groups at 3 and 5 positions. The enzyme specifically uses UDP-Glc as the sugar donor. The specificity was confirmed by expression of the 3′GT cDNA in transgenic petunia (Petunia hybrida). This is the first report of the gene isolation of a B-ring-specific glucosyltransferase of anthocyanins, which paves the way to modification of flower color by production of blue anthocyanins