研究者詳細 - 今村　維克

Impacts of Compression on Crystallization Behavior of Freeze-Dried Amorphous Sucrose 査読

Koreyoshi Imamura, Mayo Nomura, Kazuhiro Tanaka, Nobuhide Kataoka, Jun Oshitani, Hiroyuki Imanaka, Kazuhiro Nakanishi

JOURNAL OF PHARMACEUTICAL SCIENCES 99 ( 3 ) 1452 - 1463 2010年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：JOHN WILEY & SONS INC

An amorphous matrix comprised of sugar molecules is used as excipient and stabilizing agent for labile ingredients in the pharmaceutical industry. The amorphous sugar matrix is often compressed into a tablet form to reduce the volume and improve handling. Herein, the effect of compression on the crystallization behavior of an amorphous sucrose matrix was investigated. Amorphous sucrose samples were prepared by freeze-drying and compressed under different conditions, followed by analyses by differential scanning calorimetry, isothermal crystallization tests, X-ray powder diffractometry, Fourier transform infrared spectroscopy (FTIR), and gas pycnometry. The compressed sample had a lower crystallization temperature and a shorter induction period for isothermal crystallization, indicating that compression facilitates the formation of the critical nucleus of a sucrose crystal. Based on FTIR and molecular dynamics simulation results, the conformational distortion of sucrose molecules due to the compression appears to contribute to the increase in the free energy of the system, which leads to the facilitation of critical nucleus formation. An isothermal crystallization test indicated an increase in the growth rate of sucrose crystals by the compression. This can be attributed to the transformation of the microstructure from porous to nonporous, as the result of compression. (C) 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 99:1452-1463, 2010

DOI： 10.1002/jps.21890

Cultivation characteristics and gene expression profiles of Aspergillus oryzae by membrane-surface liquid culture, shaking-flask culture, and agar-plate culture 査読

Hiroyuki Imanaka, Soukichi Tanaka, Bin Feng, Koreyoshi Imamura, Kazuhiro Nakanishi

JOURNAL OF BIOSCIENCE AND BIOENGINEERING 109 ( 3 ) 267 - 273 2010年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SOC BIOSCIENCE BIOENGINEERING JAPAN

We cultivated a filamentous fungus, Aspergillus oryzae IAM 2706 by three different cultivation methods, i.e., shaking-flask culture (SFC), agar-plate culture (APC), and membrane-surface liquid culture (MSLC), to elucidate the differences of its behaviors by different cultivation methods under the same media, by measuring the growth, secretion of proteases and alpha-amylase, secreted protein level, and gene transcriptional profile by the DNA microarray analysis. The protease activities detected by MSLC and APC were much higher than that by SFC, using both modified Czapek-Dox (mCD) and dextrin-peptone-yeast extract (DPY) media. The alpha-amylase activity was detected in MSLC and APC in a much larger extent than that in SFC when DPY medium was used. On the basis of SDS-PAGE analyses and N-terminal amino acid sequences, 6 proteins were identified in the supernatants of the culture broths using DPY medium, among which oryzin (alkaline protease) and alpha-amylase were detected at a much higher extent for APC and MSLC than those for SFC while only oryzin was detected in mCD medium, in accordance with the activity measurements. A microarray analysis for the fungi cultivated by SFC, APC, and MSLC using mCD medium was carried out to elucidate the differences in the gene transcriptional profile by the cultivation methods. The gene transcriptional profile obtained for the MSLC sample showed a similar tendency to the APC sample while it was quite different from that for the SFC sample. Most of the genes specifically transcribed in the MSLC sample versus those in the SFC sample with a 10-fold up-regulation or higher were unknown or predicted proteins. However, transcription of oryzin gene was only slightly up-regulated in the MSLC sample and that of alpha-amylase gene, slightly down-regulated. (C) 2009, The Society for Biotechnology, Japan. All rights reserved.

DOI： 10.1016/j.jbiosc.2009.09.004

低温度領域における糖類アモルファスマトリクスのタンパク質安定化作用と糖分子の束縛状態

今村維克, 横山徹, 大山健一, 絹畠光倫, 今中洋行, 中西一弘

化学工学会研究発表講演要旨集 2010 584 - 584 2010年

各種糖類アモルファスマトリクスに収着した水の状態とガラス転移温度との関係

籠谷亮, 衣川耕史, 野村真世, 今中洋行, 今村維克, 中西一弘

化学工学会研究発表講演要旨集 2010 583 - 583 2010年

Streptmyces mobaraensis由来epsilon-Lysine acylaseによるN-epsilon-Lauroyl-L-Lysineの効率的合成

内田達也, 田口友造, 今中洋行, 今村維克, 是石真友子, 中西一弘

化学工学会研究発表講演要旨集 2010 1003 - 1003 2010年

クッションタンパク質を用いた機能的ペプチド固定化法の検討

國方俊暢, 今中洋行, 今村維克, 中西一弘

化学工学会研究発表講演要旨集 2010 997 - 997 2010年

オートトランスポーターを用いたペプチド表層提示系の検討

今中洋行, 植田久子, 今村維克, 中西一弘

化学工学会研究発表講演要旨集 2010 165 - 165 2010年

Development of peptide immobilization method using PS-tagged cushion protein 査読

Hiroyuki Imanaka, Daisuke Yamazumi, Toshinobu Kunikata, Koreyoshi Imamura, Kazuhiro Nakanishi

JOURNAL OF BIOSCIENCE AND BIOENGINEERING 108 S161 - S161 2009年11月

Formation characteristics of calcium phosphate deposits on a metal surface by H2O2-electrolysis reaction under various conditions 査読

Kazuaki Kanamoto, Koreyoshi Imamura, Nobuhide Kataoka, Jun Oshitani, Hiroyuki Imanaka, Kazuhiro Nakanishi

COLLOIDS AND SURFACES A-PHYSICOCHEMICAL AND ENGINEERING ASPECTS 350 ( 1-3 ) 79 - 86 2009年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER SCIENCE BV

The cathodic electrolysis of H2O2 (H2O2 + e(-) -> OH- + (OH)-O-center dot) on a metal surface in the presence of calcium and phosphate ions results in the formation of calcium phosphate deposits oil the metal surface. In this Study, the deposits formed under various treatment conditions (pHs, concentrations and ratios of calcium/phosphate ions, and so on) were characterized by scanning electron spectroscopy (SEM). and X-ray diffractometry. The exclusive formation of hydroxyapatite, HAP, was observed under comparatively narrow conditions (pH 3-4, [Ca+]/[PO43-] = 25 mM/15 mM), which is clearly different from the reported conditions for the deposition of HAP on titanium substrates. HAP was deposited in the form of a layer. comprised of morphologically amorphous HAP flakes that were less than 20 nm thick. SEM and FTIR analyses of the deposit at different stages of H2O2-electrolysis revealed that a few dozen nanometer-sized spheres of amorphous calcium phosphate were formed in the first step and then fused with each other to form ribbon-like flakes of HAP or broken glass-like brushite, depending on the pH. The pH for HAP formation oil a stainless steel surface was markedly lower than that used for titanium, and the observed process by which amorphous calcium phosphate is converted to HAP was markedly different from that for the electrochemical deposition (electrolysis of water) of HAP on a titanium substrate. (C) 2009 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.colsurfa.2009.09.007

Temperature Scanning FTIR Analysis of Secondary Structures of Proteins Embedded in Amorphous Sugar Matrix 査読

Koreyoshi Imamura, Ken-Ichi Ohyama, Toru Yokoyama, Yoshinobu Maruyama, Nakanishi Kazuhiro

JOURNAL OF PHARMACEUTICAL SCIENCES 98 ( 9 ) 3088 - 3098 2009年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：JOHN WILEY & SONS INC

Heat-induced changes in secondary structures of five proteins (bovine serum albumin, BSA; human serum albumin, HSA; myoglobin; ribonuclease A, RNase A; and, beta-lactoglobulin, beta-Lg) in an amorphous sugar matrix were analyzed by temperature-scanning Fourier transform infrared spectroscopy to elucidate the mechanism of heat-induced conformational change of solid-phase proteins. Three sugars, trehalose, maltose, and dextran (MW 6000), were used. Loss of alpha-helices due to increasing temperature was observed for BSA, HSA, and myoglobin, which are rich in a-helices. RNase A showed a marked decrease in predominant secondary structural components (beta-sheet) with increasing temperature. However, no noticeable changes in the content of secondary structures, except for a slight loss of alpha-helices, were observed for beta-Lg, which is also beta-sheet-rich. These heat-induced conformational changes were significant at temperatures above the glass transition temperature. The heat-induced conformational change in BSA dried with sugar appeared time-independent and was clearly different from that due to dehydration and from the thermal conformational change for a solution of BSA. In particular, differences in secondary structural components that increased due to loss of alpha-helices were noted. (c) 2008 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:3088-3098, 2009

DOI： 10.1002/jps.21568

Purification, Characterization, Molecular Cloning, and Expression of a New Aminoacylase from Streptomyces mobaraensis That Can Hydrolyze N-(Middle/Long)-chain-fatty-acyl-L-amino Acids as Well as N-Short-chain-acyl-L-amino acids 査読

Mayuko Koreishi, Yasuyuki Nakatani, Manami Ooi, Hiroyuki Imanaka, Koreyoshi Imamura, Kazuhiro Nakanishi

BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY 73 ( 9 ) 1940 - 1947 2009年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：TAYLOR & FRANCIS LTD

We report here on the purification, characterization, molecular cloning, and expression of a new aminoacylase, initially isolated from the supernatant of Streptomyces mobaraensis (Sm-AA). Purified wild-type Sm-AA was found to be a monomeric protein with a molecular mass of 55kDa. The cloned gene of Sm-AA contained an ORF of 1,383 bp, encoding a polypeptide of 460 amino acids. A BLAST search revealed that Sm-AA belongs to the peptidase M20 family, with identities to a hypothetical protein from Streptomyces pristinaespiralis, a putative peptidase from Streptomyces averinitilis, peptidase M20 from Frankia sp., succinyl-diaminopimelate desuccinylase from Hemophilus influenzae, and aminoacylase-1 from porcine kidney at 89, 88, 67, 29, and 25% respectively. The Sm-AA gene was subcloned into an expression vector, pSH19, and was expressed in Streptomyces lividans TK24. The amount of the recombinant Sm-AA expressed in the S. lividans cells was approximately 42-fold higher than that of Sm-AA found in the supernatant of S. mobaraensis. Sm-AA showed high hydrolytic activity towards various N-acetyl-L-amino acids and N-(middle/long)-chain-fatty-acyl-L-amino acids, with a preference for the acyl derivatives of L-Met, L-Ala, L-Cys, etc. with an optimum pH and temperature for reaction of about 7.5 and 50 degrees C (at pH 7.5).

DOI： 10.1271/bbb.90081

Efficient N epsilon-lauroyl-L-lysine production by recombinant epsilon-lysine acylase from Streptomyces mobaraensis 査読

Mayuko Koreishi, Ryoko Kawasaki, Hiroyuki Imanaka, Koreyoshi Imamura, Yasuaki Takakura, Kazuhiro Nakanishi

JOURNAL OF BIOTECHNOLOGY 141 ( 3-4 ) 160 - 165 2009年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER SCIENCE BV

epsilon-Lysine acylase from Streptomyces mobaraensis (Sm-ELA), which specifically catalyzes hydrolysis of the epsilon-amide bond in various N epsilon-acyl-L-lysines, was cloned and sequenced. The Sm-ELA gene consists of a 1617-bp open reading frame that encodes a 538-amino acid protein with a molecular mass of 55,816 Da. An NCBI protein-protein BLAST search revealed that the enzyme belongs to the YtcJ-like metal-dependent amidohydrolase family, which is further characterized as the metallo-dependent hydrolase superfamily. The Sm-ELA gene was ligated into a pUC702 vector for expression in Streptomyces lividans TK24. Expression of recombinant Sm-ELA in S. lividans was approximately 300-fold higher than that in wild-type S. mobaraensis. The recombinant Sm-ELAs from the cell-free extract and Culture supernatant were purified to homogeneity. The specific activities of the purified Sm-ELAs were 2500-2800 U/mg, which were similar to that obtained for the wild-type Sm-ELA. Using the cell-free extract of the recombinant S. lividans cells, N epsilon-lauroyl-L-lysine was synthesized from 500 mM L-lysine hydrochloride and 50, 100, or 250 mM lauric acid in an aqueous buffer Solution at 37 degrees C. The yields were close to 100% after 6 and 9 1) of reaction for 50 and 100 mM lauric acid, respectively. and 90% after 24 h for 250 mM lauric acid. (C) 2009 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.jbiotec.2009.03.008

糖類アモルファス材料の水分収着特性とタンパク質安定化作用に及ぼす高圧処理の影響

今村維克, 野村真世, 木村佳文, 伊久珠代, 絹畠光倫, 今中洋行, 中西一弘

化学工学会研究発表講演要旨集 2009 779 - 779 2009年

クッションタンパク質を利用したバイオ分子固定化技術の開発と応用

麻那古秀友, 山隅大輔, 大塚隆尚, 今中洋行, 今村維克, 中西一弘

化学工学会研究発表講演要旨集 2009 1060 - 1060 2009年

PS-tag連結タンパク質を利用したバイオ分子間相互作用解析

今中洋行, 上崎英範, 國方俊暢, 今村維克, 中西一弘

化学工学会研究発表講演要旨集 2009 934 - 934 2009年

オートトランスポーターを用いたバイオ分子表層提示系

今中洋行, 植田久子, 山下麻衣, 今村維克, 中西一弘

化学工学会研究発表講演要旨集 2009 606 - 606 2009年

糖類アモルファスマトリクスにおける糖-高分子間相互作用の形成特性

今村維克, 横山徹, 大山健一, 野村真世, 今中洋行, 中西一弘

化学工学会研究発表講演要旨集 2009 608 - 608 2009年

Recent advances in controlled immobilization of proteins onto the surface of the solid substrate and its possible application to proteomics 査読

Kazuhiro Nakanishi, Takaharu Sakiyama, Yoichi Kumada, Koreyoshi Imamura, Hiroyuki Imanaka

Current Proteomics 5 ( 3 ) 161 - 175 2008年10月

記述言語：英語

Proteome analysis plays a key role in the elucidation of the functions and applications for numerous proteins. For proteome analyses, various microplate- and microarray-based techniques have been developed by a number of researchers. Their intent was to immobilize proteins on the surface of a solid substrate in a site-directed manner while retaining structure and native biological function. In this review, we focus on recent advances in immobilization methodology for proteins/enzymes on a surface, including those using the affinity peptides screened by random peptide library systems. We also discuss applications of the affinity peptide-mediated immobilization method in fields related to proteome analysis, particularly our recent work concerning immunoassay and protein-protein interaction analysis. ©2008 Bentham Science Publishers Ltd.

DOI： 10.2174/157016408785909622

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