Overexpression and purification of the recombinant Ca2+-binding protein, apoaequorin.

Abstract:

:The small, monomeric Ca2+-binding photoprotein, aequorin, emits blue light by an intramolecular reaction when mixed with Ca2+. The photoprotein is made up of coelenterazine and molecular oxygen, bound noncovalently to apoaequorin (apoprotein). The chemical steps leading to light emission, involving the oxidative degradation of coelenterazine, have been studied extensively, but little is known about the active site and how the molecule catalyzes the oxidation of coelenterazine. The three-dimensional structure of the protein has not been determined and therefore answers to these questions have remained unavailable. The present paper describes a procedure for preparing fairly large amounts of apoaequorin and aequorin for X-ray crystallographic studies. It consists of fusing the apoaequorin cDNA to the signal peptide coding sequence of the outer membrane protein A of Escherichia coli, which is under the control of the lipoprotein promoter. When the cDNA was expressed in E. coli, a large excess of the recombinant protein was produced and released into the culture medium. Purification of the protein was accomplished by acid precipitation and DEAE-cellulose chromatography. The procedure yielded 7.4 mg of recombinant apoaequorin with a purity greater than 95% from 200 ml of culture medium. On regeneration with coelenterazine, the recombinant aequorin was fully active with Ca2+.

journal_name

J Biochem

journal_title

Journal of biochemistry

authors

Inouye S,Aoyama S,Miyata T,Tsuji FI,Sakaki Y

doi

10.1093/oxfordjournals.jbchem.a122689

subject

Has Abstract

pub_date

1989-03-01 00:00:00

pages

473-7

issue

3

eissn

0021-924X

issn

1756-2651

journal_volume

105

pub_type

杂志文章
  • c-Abl is involved in the F-actin assembly triggered by L-selectin crosslinking.

    abstract::L-selectin is a cell adhesion molecule mediating the initial capture and subsequent rolling of leukocytes along the endothelial cells expressing L-selectin ligands. In addition to its action in adhesion, an intracellular signaling role for L-selectin has been recognized. Its cytoplasmic domain is involved in signal tr...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/jb/mvj149

    authors: Chen C,Ba X,Xu T,Cui L,Hao S,Zeng X

    更新日期:2006-08-01 00:00:00

  • HPC-1/syntaxin 1A suppresses exocytosis of PC12 cells.

    abstract::The membrane protein syntaxin (originally named HPC-1) is involved in vesicle trafficking and required for neurotransmitter release at nerve terminals. The presence of syntaxin on target membranes is hypothesized to confer specificity to targeting and fusion via interactions with complementary vesicle-associated prote...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a022337

    authors: Watanabe T,Fujiwara T,Komazaki S,Yamaguchi K,Tajima O,Akagawa K

    更新日期:1999-04-01 00:00:00

  • Structures of galactose-containing oligosaccharides of alpha-mannosidase from porcine kidney.

    abstract::Oligosaccharides of a non-oligomannoside type were released from porcine alpha-mannosidase by hydrazinolysis, and were fractionated into at least 15 homogeneous oligosaccharides. Most of them are oligosaccharides with galactose and N-acetylglucosamine residues attached to a common core, alpha Man2 beta Man beta GlcNAc...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a121740

    authors: Kozutsumi Y,Kawasaki T,Yamashina I

    更新日期:1986-08-01 00:00:00

  • Enzyme-linked immunoassay. I. Novel method for synthesis of the insulin-beta-D-galactosidase conjugate and its applicability for insulin assay.

    abstract::Pork insulin was subjected to mercaptosuccinylation and then coupled to beta-D-galactosidase [EC 3.2.1.23] from Escherichia coli using N,N'-o-phenylenedimaleimide. The competitive binding of the conjugate and insulin to anti-insulin antibody was tested. Results showed that formation of an insulin-beta-D-galactosidase ...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:

    authors: Kato K,Hamaguchi Y,Fukui H,Ishikawa E

    更新日期:1975-07-01 00:00:00

  • Overexpression and functional characterization of a serine carboxypeptidase inhibitor (I(C)) from Saccharomyces cerevisiae.

    abstract::Carboxypeptidase Y (CPY) inhibitor, I(C), a cytoplasmic inhibitor of vacuolar proteinases in yeast, Saccharomyces cerevisiae, was purified by means of a high-level expression system using a proteinase-deficient strain, BJ2168, and an expression vector with the promoter GAL1. The purified I(C) exists as a monomeric bet...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a003311

    authors: Mima J,Suzuki H,Takahashi M,Hayashi R

    更新日期:2002-12-01 00:00:00

  • Evidence that arachidonic acid is deficient in phosphatidylinositol of Drosophila heads.

    abstract::We have found that arachidonic (20 : 4) acid is indetectable in phosphatidylinositol and diacyglycerol extracted from Drosophila heads. After careful examinations of the lipid extraction processes and fatty acid detection system (gas-liquid chromatography), we excluded the possibility of the oxidation of polyunsaturat...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a135322

    authors: Yoshioka T,Inoue H,Kasama T,Seyama Y,Nakashima S,Nozawa Y,Hotta Y

    更新日期:1985-09-01 00:00:00

  • Role of 17-kDa essential light chain isoforms of aorta smooth muscle myosin.

    abstract::Aorta smooth muscle myosin contains two kinds of 17-kDa essential light chain, LC17nm (nonmuscle-type) and LC17gi (gizzard-type) [Hasegawa, Y., Ueda, Y., Watanabe, M., & Morita, F. (1992) J. Biochem. 111, 798-803]. The LC17 isoforms were released from porcine aorta myosin by incubation at 46 degrees C. The rate of rel...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a123840

    authors: Hasegawa Y,Morita F

    更新日期:1992-06-01 00:00:00

  • The importance of parathyroid hormone in inhibition of collagen synthesis and mitogenesis of osteoblastic cell.

    abstract::Loss of bone substance is a common manifestation of hyperparathyroidism. This suggests that parathyroid hormone (PTH) plays an important role as to bone mass. To investigate the mechanism underlying this change in bone mass, I studied the effects of PTH on collagen synthesis and mitogenesis of UMR-106 rat osteoblastic...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a122970

    authors: Pun KK

    更新日期:1989-12-01 00:00:00

  • Proteasome inhibitors block Ras/ERK signaling pathway resulting in the downregulation of Fas ligand expression during activation-induced cell death in T cells.

    abstract::Activation-induced cell death (AICD) plays a critical role in the maintenance of homeostasis and peripheral tolerance in the immune system, and is mediated by Fas ligand (FasL) expression and the interaction between Fas and FasL. In the present study, we examined the role of the ubiquitin-proteasome system in AICD usi...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a003106

    authors: Tanimoto Y,Kizaki H

    更新日期:2002-03-01 00:00:00

  • Role of Nrf2 and p62/ZIP in the neurite outgrowth by carnosic acid in PC12h cells.

    abstract::Neurotrophins such as NGF promote neuronal survival and differentiation via the cell surface TrkA neurotrophin receptor. Compounds with neurotrophic actions that are low in molecular weight and can permeate the blood-brain barrier are promising therapeutic agents against neurodegenerative diseases such as Alzheimer's ...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/jb/mvp149

    authors: Kosaka K,Mimura J,Itoh K,Satoh T,Shimojo Y,Kitajima C,Maruyama A,Yamamoto M,Shirasawa T

    更新日期:2010-01-01 00:00:00

  • Methyl hydroperoxy-epoxy-octadecenoate as an autoxidation product of methyl linoleate: a new inhibitor-uncoupler of mitochondrial respiration.

    abstract::A new peroxide compound (ML-X) was isolated from an autoxidation product of methyl linoleate and was determined as methyl 9-hydroperoxy-12, 13-epoxy-10-octadecenoate. This compound inhibited state 3 respiration of rat heart- and liver mitochondria when glutamate and malate were used as substrates, but not when the sub...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a134027

    authors: Imagawa T,Kasai S,Matsui K,Nakamura T

    更新日期:1982-10-01 00:00:00

  • Hepatocyte growth factor suppresses the onset of liver cirrhosis and abrogates lethal hepatic dysfunction in rats.

    abstract::Heptic fibrosis/cirrhosis is a common hepatic disease characterized by the hyper-accumulation of connective tissue components, and hepatic necrosis. Chronic alcohol ingestion, viral infection, and metabolic disorders are contributing factors and there has been no effective treatment. Hepatocyte growth factor (HGF), or...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a124958

    authors: Matsuda Y,Matsumoto K,Ichida T,Nakamura T

    更新日期:1995-09-01 00:00:00

  • Biochemical characterization of galloyl pedunculagin (ellagitannin) as a selective inhibitor of the beta-regulatory subunit of A-kinase in vitro.

    abstract::The inhibitory effects of galloyl pedunculagin (GP) and eugeniin on the phosphorylation of histone H2B by cAMP-dependent protein kinase (A-kinase) and autophosphorylation of its beta-regulatory subunit (A-kinase beta) were examined in vitro. It was found that (i) GP (ID(50) = approx. 50 nM) effectively inhibits the ac...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a002871

    authors: Kosuge S,Maekawa T,Saito C,Tanaka T,Kouno I,Ohtsuki K

    更新日期:2001-03-01 00:00:00

  • Mechanism-based inactivation of coenzyme B12-dependent diol dehydratase by 3-unsaturated 1,2-diols and thioglycerol.

    abstract::The reactions of diol dehydratase with 3-unsaturated 1,2-diols and thioglycerol were investigated. Holodiol dehydratase underwent rapid and irreversible inactivation by either 3-butene-1,2-diol, 3-butyne-1,2-diol or thioglycerol without catalytic turnovers. In the inactivation, the Co-C bond of adenosylcobalamin under...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/jb/mvn086

    authors: Toraya T,Tamura N,Watanabe T,Yamanishi M,Hieda N,Mori K

    更新日期:2008-10-01 00:00:00

  • Enhancement of the mitogenic effect by artificial juxtacrine stimulation using immobilized EGF.

    abstract::Mouse epidermal growth factor (EGF) was covalently conjugated with the water-soluble polymer, poly(acrylic acid) (EGF-PAA), or with the water-insoluble polymer, surface-hydrolyzed poly(methyl methacrylate) (EGF-PMMA). Immobilized EGF (EGF-PMMA) stimulated DNA synthesis in Chinese hamster ovary cells overexpressing EGF...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a021616

    authors: Ito Y,Li JS,Takahashi T,Imanishi Y,Okabayashi Y,Kido Y,Kasuga M

    更新日期:1997-03-01 00:00:00

  • A novel method to detect translation of membrane proteins following microvesicle intercellular transfer of nucleic acids.

    abstract::Microvesicles (MVs) serve as vectors of nucleic-acid dissemination and are important mediators of intercellular communication. However, the functionality of packaged nucleic acids on recipient cells following transfer of MV cargo has not been clearly elucidated. This limitation is attributed to a lack of methodology a...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/jb/mvw033

    authors: Lu JF,Pokharel D,Padula MP,Bebawy M

    更新日期:2016-11-01 00:00:00

  • The basigin group of the immunoglobulin superfamily: complete conservation of a segment in and around transmembrane domains of human and mouse basigin and chicken HT7 antigen.

    abstract::Basigin belongs to the immunoglobulin superfamily and may be related to the primordial form of the superfamily. Human basigin cDNA was isolated and sequenced, and the predicted protein structure was compared with that of mouse basigin and two related molecules, embigin and the chicken blood-brain barrier antigen HT7. ...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a123657

    authors: Miyauchi T,Masuzawa Y,Muramatsu T

    更新日期:1991-11-01 00:00:00

  • Uncoupling of ATP splitting from Ca(2+)-transport in Ca(2+)-transporting ATPase of the sarcoplasmic reticulum as a result of modification by N-(3-pyrene)maleimide: activation of a channel with a specificity for alkaline earth metal ions.

    abstract::The sarcoplasmic reticulum (SR) membranes of rabbit skeletal muscle were allowed to react with N-(3-pyrene)maleimide (PMI) at pH 7 at 30 degrees C. The Ca(2+)-transporting activity of the SR membranes was reduced to 20% when PMI was bound to the extent of 1 mol/mol of Ca(2+)-transporting ATPase. The ATPase and the E-P...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a124155

    authors: Suzuki T,Kawakita M

    更新日期:1993-08-01 00:00:00

  • Structure of mouse interferon stimulated gene factor 3 gamma (ISGF3 gamma/p48) cDNA and chromosomal localization of the gene.

    abstract::Interferon stimulated gene factor 3 (ISGF3) is a trimeric transcription factor activated on treatment of cells with interferon-alpha and beta (type I IFNs). Upon stimulation, the regulatory subunits, p84/91 and p113, present in the cytoplasm are phosphorylated at specific tyrosine residues and assemble with the DNA bi...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a021227

    authors: Suhara W,Yoneyama M,Yonekawa H,Fujita T

    更新日期:1996-02-01 00:00:00

  • Modification of glucoamylases from Rhizopus sp. with 1-cyclohexyl-3-(2-morpholinyl-(4)-ethyl)carbodiimide.

    abstract::To investigate the role of carboxyl groups of glucoamylases [EC 3.2.1.3] from a Rhizopus sp. (Gluc1 and Gluc2), the modification of Gluc1 and Gluc2 with a water-soluble carbodiimide, 1-cyclohexyl-3-(2-morpholinyl-(4)-ethyl)carbodiimide metho-p-toluenesulfonate (CMC), was studied. The inactivation of Gluc1 proceeded wi...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a134842

    authors: Iwama M,Ohtsuki R,Takahashi T,Irie M

    更新日期:1984-08-01 00:00:00

  • Light-induced pH changes and changes in absorbance of pH indicators in Rhodospirillum rubrum chromatophores.

    abstract::1. The light-induced pH change of chromatophore suspensions from Rhodospirillum rubrum was stimulated significantly and similarly by KCl, NaCl, LiCl, RbCl, CsCl, MgCl2, MnCl2, and CaCl2. In the dark, the pH of chromatophore suspensions decreased immediately and markedly on adding these salts. 2. The light-induced pH c...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a131814

    authors: Nishi N,Sakata-Sogawa K,Soe G,Yamashita J

    更新日期:1977-11-01 00:00:00

  • Isolation and characterization of NADP+-specific isocitrate dehydrogenase from the pupa of Bombyx mori.

    abstract::NADP+-specific isocitrate dehydrogenase was found in several tissues of the pupa of the silkworm, Bombyx mori. This enzyme was highly purified from the whole bodies of pupae. This is the first isolation of the enzyme from insect materials. The purified enzyme gave a single protein band on polyacrylamide gel electropho...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:

    authors: Miake F,Torikata T,Koga K,Hayashi K

    更新日期:1977-08-01 00:00:00

  • Partial purification of trimethylamine-N-oxide (TMAO) demethylase from crude fish muscle microsomes by detergents.

    abstract::Detergent treatments were examined for their efficacy in purifying trimethylamine-N-oxide (TMAO) demethylase activity from fish muscle microsomes. Tritons X-100 and X-45, deoxycholate, Brijs, Tweens 20, 65, and 80, and SDS were generally ineffective in solubilizing demethylase activity from this membrane fraction, at ...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a121709

    authors: Parkin KL,Hultin HO

    更新日期:1986-07-01 00:00:00

  • Characterization of a novel endo-type alginate lyase derived from Shewanella sp. YH1.

    abstract::Alginate, which is an anionic polysaccharide, is widely distributed in the cell wall of brown algae. Alginate and the products of its degradation (oligosaccharides) are used in stabilizers, thickeners and gelling agents, especially in the food industry. The degradation of alginate generally involves a combination of s...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/jb/mvy001

    authors: Yagi H,Fujise A,Itabashi N,Ohshiro T

    更新日期:2018-04-01 00:00:00

  • Store depletion by caffeine/ryanodine activates capacitative Ca(2+) entry in nonexcitable A549 cells.

    abstract::Capacitative Ca(2+) entry is essential for refilling intracellular Ca(2+) stores and is thought to be regulated primarily by inositol 1, 4,5-trisphosphate (IP(3))-sensitive stores in nonexcitable cells. In nonexcitable A549 cells, the application of caffeine or ryanodine induces Ca(2+) release in the absence of extrac...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a022757

    authors: Xue HH,Zhao DM,Suda T,Uchida C,Oda T,Chida K,Ichiyama A,Nakamura H

    更新日期:2000-08-01 00:00:00

  • Dinuclear lanthanum(III) complex for efficient hydrolysis of RNA.

    abstract::A dinuclear La3+ complex hydrolyzes the phosphodiester bond in diribonucleotides efficiently under mild conditions. The rate of hydrolysis is remarkably accelerated on the complex formation between La3+ and TPHP (TPHP = N,N,N',N'-tetrakis [(2-pyridyl)methyl]-2-hydroxy-1,3-diaminopropane), and the highest activity is a...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a021521

    authors: Yashiro M,Ishikubo A,Komiyama M

    更新日期:1996-12-01 00:00:00

  • Gene structure and multiple mRNA species of Drosophila melanogaster aldolase generating three isozymes with different enzymatic properties.

    abstract::Genomic clones encoding the Drosophila aldolase gene were isolated and the organization of the gene was determined. The protein-coding region spanning nearly 3.5 kb consists of five coding exons (exon 2, 3, 4 alpha, 4 beta, and 4 gamma). The insect exon 2 corresponds to exons 2 to 7 of vertebrate aldolase genes and th...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a123958

    authors: Kai T,Sugimoto Y,Kusakabe T,Zhang R,Koga K,Hori K

    更新日期:1992-11-01 00:00:00

  • Purification and some properties of chalcone synthase from a carrot suspension culture induced for anthocyanin synthesis and preparation of its specific antiserum.

    abstract::Chalcone synthase was purified to homogeneity by polyacrylamide gel electrophoresis from cell suspension cultures of carrot in which anthocyanin synthesis was induced by transferring the cells from a medium containing 2,4-dichlorophenoxy-acetic acid (2,4-D) to one lacking it. A molecular weight of 80,000-85,000 for th...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a135278

    authors: Ozeki Y,Sakano K,Komamine A,Tanaka Y,Noguchi H,Sankawa U,Suzuki T

    更新日期:1985-07-01 00:00:00

  • Characterization of novel acetyltransferases found in budding and fission yeasts that detoxify a proline analogue, azetidine-2-carboxylic acid.

    abstract::We recently found that budding yeast Saccharomyces cerevisiae sigma1278b, but not genome project strain S288C, has a gene conferring resistance to L-azetidine-2-carboxylic acid (AZC), a toxic four-membered ring analogue of L-proline. Also, the gene, designated as MPR1, encodes a novel acetyltransferase that detoxifies...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/jb/mvg003

    authors: Nomura M,Nakamori S,Takagi H

    更新日期:2003-01-01 00:00:00

  • Binding of activated macrophages to tumor cells through a macrophage lectin and its role in macrophage tumoricidal activity.

    abstract::A 45-60 kDa Gal/GalNAc-specific macrophage lectin was found to participate in the interaction between tumor cells and tumoricidal macrophages activated by an antitumor streptococcal preparation, OK-432, and in the tumoricidal activity of the activated macrophages. The binding between OK-432-elicited activated macropha...

    journal_title:Journal of biochemistry

    pub_type: 杂志文章

    doi:10.1093/oxfordjournals.jbchem.a122763

    authors: Oda S,Sato M,Toyoshima S,Osawa T

    更新日期:1989-06-01 00:00:00