A base pair at the bottom of the anticodon stem is reciprocally preferred for discrimination of cognate tRNAs by Escherichia coli lysyl- and glutaminyl-tRNA synthetases

Although the yeast amber suppressor tRNA(Tyr) is a good candidate for a carrier of unnatural amino acids into proteins, slight misacylation with lysine was found to occur in an Escherichia coli protein synthesis system. Although it was possible to restrain the mislysylation by genetically engineering the anticodon stem region of the amber suppressor tRNA(Tyr), the mutant tRNA showing the lowest acceptance of lysine was found to accept a trace level of glutamine instead. Moreover, the glutamine-acceptance of various tRNA(Tyr) transcripts substituted at the anticodon stem region varied in reverse proportion to the lysine-acceptance, similar to a ‘seesaw’. The introduction of a C31–G39 base pair at the site was most effective for decreasing the lysine-acceptance and increasing the glutamine-acceptance. When the same substitution was introduced into E.coli tRNA(Lys) transcripts, the lysine-accepting activity was decreased by 100-fold and faint acceptance of glutamine was observed. These results may support the idea that there are some structural element(s) in the anticodon stem of tRNA, which are not shared by aminoacyl-tRNA synthetases that have similar recognition sites in the anticodon, such as E.coli lysyl- and glutaminyl-tRNA synthetases

Structural basis for recognition of cognate tRNA by tyrosyl-tRNA synthetase from three kingdoms

The specific aminoacylation of tRNA by tyrosyl-tRNA synthetases (TyrRSs) relies on the identity determinants in the cognate tRNATyrs. We have determined the crystal structure of Saccharomyces cerevisiae TyrRS (SceTyrRS) complexed with a Tyr-AMP analog and the native tRNATyr(GΨA). Structural information for TyrRS–tRNATyr complexes is now full-line for three kingdoms. Because the archaeal/eukaryotic TyrRSs–tRNATyrs pairs do not cross-react with their bacterial counterparts, the recognition modes of the identity determinants by the archaeal/eukaryotic TyrRSs were expected to be similar to each other but different from that by the bacterial TyrRSs. Interestingly, however, the tRNATyr recognition modes of SceTyrRS have both similarities and differences compared with those in the archaeal TyrRS: the recognition of the C1-G72 base pair by SceTyrRS is similar to that by the archaeal TyrRS, whereas the recognition of the A73 by SceTyrRS is different from that by the archaeal TyrRS but similar to that by the bacterial TyrRS. Thus, the lack of cross-reactivity between archaeal/eukaryotic and bacterial TyrRS-tRNATyr pairs most probably lies in the different sequence of the last base pair of the acceptor stem (C1-G72 vs G1-C72) of tRNATyr. On the other hand, the recognition mode of Tyr-AMP is conserved among the TyrRSs from the three kingdoms

ジドウ ヨウゴ シセツ ニケル ショクイン ノ シシツ ノ コウジョウ エノ トリクミ ダイニホウ ジドウ ヨウゴ シセツ ニオケル フクシ ノ ジンザイ イクセイ

今回は施設実習を行なう実習生への意識調査を通して実習前と後の意識の違い、子どもへの援助のあり方、職員へのイメージ、反省会での関わり方を調査した。実習前の暗いイメージが実習後には明るくなっている。又、職員に対しても優しいとか温かいイメージに変化している。実習そのものでは、辛いという意識、けんかなどへの対応の不安を持っていた。しかし、自由記述では、施設への就職希望が見られた。また、実習を通して自分自身の生き方を見直している。実習から学習意欲を持った学生もみられた。大学での講義やオリエンテーションにて子どもへの具体的な対応や施設説明がより学生に理解できる内容の連携が求められる。施設の課題として実習生に対してさらに効果的な実習プログラムの見直しの必要がある

児童福祉施設における職員の資質の向上への取組み「第一報」 : M学園の現状と課題を通して児童養護の今後を探る

最近の乳幼児を取り巻く環境は「子どもにとって幸せ」という状態では決して言えない。大人が子どもを殺したり、虐待したりする悲惨な事件が日本中で起きている。そのような中で福祉施設の果たす役割は重要である。しかし、子どもを預ける保護者が施設に対するイメージの持ち方は、預ける前はマイナスのイメージであったが入所後は良いイメージを持つようになった。それは、子どもが入所したことで少なくとも「良い方向」に変化していることであった。しかし、施設の関係者と医者など専門家との連携の困難さや重要さも理解できた。また、職員の指導のあり方で子どもは「生きている喜び」や「生きていたい」と言う意欲をもつことができる。M学園の運営方針や事例を検討したことで福祉施設として職員の指導の現状と課題が理解できた。さらに、保護者の意識調査を通して今後どのように保護者に対して対応するか見直す視点となった

A Keplerian Circumbinary Disk around the Protobinary System L1551 NE

We present SubMillimeter-Array observations of a Keplerian disk around the Class I protobinary system L1551 NE in 335 GHz continuum emission and submillimeter line emission in 13CO (J=3-2) and C18O (J=3-2) at a resolution of ~120 x 80 AU. The 335-GHz dust-continuum image shows a strong central peak closely coincident with the binary protostars and likely corresponding to circumstellar disks, surrounded by a ~600 x 300 AU feature elongated approximately perpendicular to the [Fe II] jet from the southern protostellar component suggestive of a circumbinary disk. The 13CO and C18O images confirm that the circumbinary continuum feature is indeed a rotating disk; furthermore, the C18O channel maps can be well modeled by a geometrically-thin disk exhibiting Keplerian rotation. We estimate a mass for the circumbinary disk of ~0.03-0.12 Msun, compared with an enclosed mass of ~0.8 Msun that is dominated by the protobinary system. Compared with several other Class I protostars known to exhibit Keplerian disks, L1551 NE has the lowest bolometric temperature (~91 K), highest envelope mass (~0.39 Msun), and the lowest ratio in stellar mass to envelope + disk + stellar mass (~0.65). L1551 NE may therefore be the youngest protostellar object so far found to exhibit a Keplerian disk. Our observations present firm evidence that Keplerian disks around binary protostellar systems, ``Keplerian circumbinary disks', can exist. We speculate that tidal effects from binary companions could transport angular momenta toward the inner edge of the circumbinary disk and create the Keplerian circumbinary disk.Comment: 27 pages, 8 figure

Characterization and tRNA Recognition of Mammalian Mitochondrial Seryl-tRNA Synthetase

Animal mitochondrial protein synthesis systems contain two serine tRNAs (tRNAs(Ser)) corresponding to the codons AGY and UCN, each possessing an unusual secondary structure; the former lacks the entire D arm, and the latter has a slightly different cloverleaf structure. To elucidate whether these two tRNAs(Ser) can be recognized by the single animal mitochondrial seryl-tRNA synthetase (mt SerRS), we purified mt SerRS from bovine liver 2400-fold and showed that it can aminoacylate both of them. Specific interaction between mt SerRS and either of the tRNAs(Ser) was also observed in a gel retardation assay. cDNA cloning of bovine mt SerRS revealed that the deduced amino acid sequence of the enzyme contains 518 amino acid residues. The cDNAs of human and mouse mt SerRS were obtained by reverse transcription-polymerase chain reaction and expressed sequence tag data base searches. Elaborate inspection of primary sequences of mammalian mt SerRSs revealed diversity in the N-terminal domain responsible for tRNA recognition, indicating that the recognition mechanism of mammalian mt SerRS differs considerably from that of its prokaryotic counterpart. In addition, the human mt SerRS gene was found to be located on chromosome 19q13.1, to which the autosomal deafness locus DFNA4 is mapped

Implantation of Liquid Nitrogen Frozen Tumor Tissue after Posterior Decompression and Stabilization for Metastatic Spinal Tumors

Study DesignA retrospective study.PurposeTo evaluate the immunity-enhancing effect of implantation of a liquid nitrogen-treated tumor.Overview of LiteratureWe have developed a new technique of implanting a tumor frozen in liquid nitrogen after posterior decompression and stabilization, with the aim of enhancing antitumor immunity in order to prolong the survival period of the patient. In the current study, the immunity-enhancing effect of this new technique has been evaluated.MethodsThe subjects were 19 patients in whom we had earlier performed decompression and stabilization between April 2011 and September 2013. The 19 subjects were divided into two groups, namely a frozen autologous tumor tissue implantation group (n=15; "implantation group"), which consisted of patients, who underwent implantation with autologous tumor tissue frozen in liquid nitrogen, and a control group (n=4), which consisted of patients, who did not undergo autologous cancer transplantation. To evaluate the immunity-enhancing effect of the protocol, plasma cytokines (interferon [IFN]-γ and interleukin [IL]-12) were analyzed before surgery and a month after surgery.ResultsThe mean rate of increase in IFN-γ was significantly higher in the implantation group (p=0.03). Regarding IL-12, no significant difference was observed between the groups, although the implantation group exhibited increased levels of IL-12 (p=0.22).ConclusionsDecompression and stabilization combined with autologous frozen tumor cell implantation can enhance cancer immunity in metastatic spinal tumor patients. It is hypothesized that this procedure might prevent local recurrence and prolong survival period

Surgical site infection after total en bloc spondylectomy: Risk factors and the preventive new technology

Background context Surgical site infection (SSI) associated with instruments remains a serious and common complication in patients who undergo total en bloc spondylectomy (TES). It is very important that the risk factors for SSI are known to prevent it.Purpose The purpose of the study was to identify independent risk factors for SSI after TES and evaluate the positive effect of iodine-supported spinal instruments in the prevention of SSI after TES.Study design This is a retrospective clinical study.Patient sample One hundred twenty-five patients who underwent TES for vertebral tumor were evaluated.Outcome measures Incidence rate of SSI, risk factors for SSI after TES, and safety of iodine-supported spinal instruments were the outcome measures.Methods Risk factors for SSI were analyzed using logistic regression. In recent 69 patients with iodine-supported spinal instruments, the thyroid hormone levels in the blood were examined to confirm if iodine from the implant influenced thyroid function. Postoperative radiological evaluations were performed regularly.Results The rate of SSI was 6.4% (8/125 patients). By multivariate logistic regression, combined anterior and posterior approach and nonuse of iodine-supported spinal instruments were associated with an increased risk of SSI. The rate of SSI without iodine-supported spinal instruments was 12.5%, whereas the rate with iodine-supported spinal instruments was 1.4%. This difference was statistically significant. There were no detected abnormalities of thyroid gland function with the use of iodine-supported instruments. Among the 69 patients with iodine-supported spinal instruments, 2 patients required additional surgery because of instrument failure. However, there were no obvious involvements with the use of iodine-supported spinal instruments.Conclusions This study identified combined anterior and posterior approach and nonuse of iodine-supported spinal instruments to be independent risk factors for SSI after TES. Iodine-supported spinal instrument was extremely effective for prevention of SSI in patients with compromised status, and it had no detection of cytotoxic or adverse effects on the patients

Functional replacement of the endogenous tyrosyl-tRNA synthetase–tRNATyr pair by the archaeal tyrosine pair in Escherichia coli for genetic code expansion

Non-natural amino acids have been genetically encoded in living cells, using aminoacyl-tRNA synthetase–tRNA pairs orthogonal to the host translation system. In the present study, we engineered Escherichia coli cells with a translation system orthogonal to the E. coli tyrosyl-tRNA synthetase (TyrRS)–tRNATyr pair, to use E. coli TyrRS variants for non-natural amino acids in the cells without interfering with tyrosine incorporation. We showed that the E. coli TyrRS–tRNATyr pair can be functionally replaced by the Methanocaldococcus jannaschii and Saccharomyces cerevisiae tyrosine pairs, which do not cross-react with E. coli TyrRS or tRNATyr. The endogenous TyrRS and tRNATyr genes were then removed from the chromosome of the E. coli cells expressing the archaeal TyrRS–tRNATyr pair. In this engineered strain, 3-iodo-l-tyrosine and 3-azido-l-tyrosine were each successfully encoded with the amber codon, using the E. coli amber suppressor tRNATyr and a TyrRS variant, which was previously developed for 3-iodo-l-tyrosine and was also found to recognize 3-azido-l-tyrosine. The structural basis for the 3-azido-l-tyrosine recognition was revealed by X-ray crystallography. The present engineering allows E. coli TyrRS variants for non-natural amino acids to be developed in E. coli, for use in both eukaryotic and bacterial cells for genetic code expansion