Publikation:

General mutagenesis/gene expression procedure for the construction of variant immunoglobulin domains in Escherichia coli : Production of the Bence-Jones protein REIv via fusion to beta-lactamase

Lade...
Vorschaubild

Dateien

Darom_222146_MD in KOPS edit.pdf
Darom_222146_MD in KOPS edit.pdfGröße: 2.53 MBDownloads: 433

Datum

1992

Autor:innen

Kolmar, Harald
Hennecke, Frank
Wippler, Jürgen
Fritz, Hans-Joachim

Herausgeber:innen

Kontakt

ISSN der Zeitschrift

Electronic ISSN

ISBN

Bibliografische Daten

Verlag

Schriftenreihe

Auflagebezeichnung

ArXiv-ID

Internationale Patentnummer

Angaben zur Forschungsförderung

Projekt

Open Access-Veröffentlichung
Open Access Green
Core Facility der Universität Konstanz

Gesperrt bis

Titel in einer weiteren Sprache

Publikationstyp
Zeitschriftenartikel
Publikationsstatus
Published

Erschienen in

Journal of Molecular Biology. 1992, 228(2), pp. 359-365. ISSN 0022-2836. Available under: doi: 10.1016/0022-2836(92)90826-6

Zusammenfassung

A novel mutagenesis/gene expression and protein purification scheme was established for ready construction and purification of variant immunoglobulin domains in Escherichia coli. This procedure, which has been applied to the production of the Vκ domain of the Bence-Jones protein REI and structural variants of it, rests on the synthesis of chimeric proteins with β-lactamase as the amino-terminal fusion partner. The β-lactamase not only guides the fusion protein to the periplasmic space, but also allows affinity chromatography on phenylboronate-Sepharose as an efficient and general purification procedure, independent of hypervariable loop structure. The REIv protein was released from the purified fusion protein by site-specific proteolytic cleavage. After a second passage through the same affinity column, up to 2 mg of pure REIv was obtained starting from one liter of bacterial liquid culture. A scheme of oligonucleotide-directed mutagenesis was introduced for replacement of DNA stretches encoding hypervariable loops. It exploits a colony color genetic screen and can be applied to any DNA sequence replacement. Mutations can be constructed by simple co-transformation with single-stranded template DNA and mutagenic oligonucleotide.

Zusammenfassung in einer weiteren Sprache

Fachgebiet (DDC)
570 Biowissenschaften, Biologie

Schlagwörter

Konferenz

Rezension
undefined / . - undefined, undefined

Forschungsvorhaben

Organisationseinheiten

Zeitschriftenheft

Zugehörige Datensätze in KOPS

Zitieren

ISO 690KOLMAR, Harald, Elisa FERRANDO-MAY, Frank HENNECKE, Jürgen WIPPLER, Hans-Joachim FRITZ, 1992. General mutagenesis/gene expression procedure for the construction of variant immunoglobulin domains in Escherichia coli : Production of the Bence-Jones protein REIv via fusion to beta-lactamase. In: Journal of Molecular Biology. 1992, 228(2), pp. 359-365. ISSN 0022-2836. Available under: doi: 10.1016/0022-2836(92)90826-6
BibTex
@article{Kolmar1992Gener-22065,
  year={1992},
  doi={10.1016/0022-2836(92)90826-6},
  title={General mutagenesis/gene expression procedure for the construction of variant immunoglobulin domains in Escherichia coli : Production of the Bence-Jones protein REIv via fusion to beta-lactamase},
  number={2},
  volume={228},
  issn={0022-2836},
  journal={Journal of Molecular Biology},
  pages={359--365},
  author={Kolmar, Harald and Ferrando-May, Elisa and Hennecke, Frank and Wippler, Jürgen and Fritz, Hans-Joachim}
}
RDF
<rdf:RDF
    xmlns:dcterms="http://purl.org/dc/terms/"
    xmlns:dc="http://purl.org/dc/elements/1.1/"
    xmlns:rdf="http://www.w3.org/1999/02/22-rdf-syntax-ns#"
    xmlns:bibo="http://purl.org/ontology/bibo/"
    xmlns:dspace="http://digital-repositories.org/ontologies/dspace/0.1.0#"
    xmlns:foaf="http://xmlns.com/foaf/0.1/"
    xmlns:void="http://rdfs.org/ns/void#"
    xmlns:xsd="http://www.w3.org/2001/XMLSchema#" > 
  <rdf:Description rdf:about="https://kops.uni-konstanz.de/server/rdf/resource/123456789/22065">
    <dspace:hasBitstream rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/22065/2/Darom_222146_MD%20in%20KOPS%20edit.pdf"/>
    <dcterms:bibliographicCitation>Journal of molecular biology ; 228 (1992), 2. - S. 359-365</dcterms:bibliographicCitation>
    <foaf:homepage rdf:resource="http://localhost:8080/"/>
    <dcterms:issued>1992</dcterms:issued>
    <dcterms:abstract xml:lang="eng">A novel mutagenesis/gene expression and protein purification scheme was established for ready construction and purification of variant immunoglobulin domains in Escherichia coli. This procedure, which has been applied to the production of the Vκ domain of the Bence-Jones protein REI and structural variants of it, rests on the synthesis of chimeric proteins with β-lactamase as the amino-terminal fusion partner. The β-lactamase not only guides the fusion protein to the periplasmic space, but also allows affinity chromatography on phenylboronate-Sepharose as an efficient and general purification procedure, independent of hypervariable loop structure. The REIv protein was released from the purified fusion protein by site-specific proteolytic cleavage. After a second passage through the same affinity column, up to 2 mg of pure REIv was obtained starting from one liter of bacterial liquid culture. A scheme of oligonucleotide-directed mutagenesis was introduced for replacement of DNA stretches encoding hypervariable loops. It exploits a colony color genetic screen and can be applied to any DNA sequence replacement. Mutations can be constructed by simple co-transformation with single-stranded template DNA and mutagenic oligonucleotide.</dcterms:abstract>
    <dc:language>eng</dc:language>
    <dc:creator>Hennecke, Frank</dc:creator>
    <dc:contributor>Wippler, Jürgen</dc:contributor>
    <dspace:isPartOfCollection rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/28"/>
    <dc:creator>Fritz, Hans-Joachim</dc:creator>
    <dcterms:isPartOf rdf:resource="https://kops.uni-konstanz.de/server/rdf/resource/123456789/28"/>
    <dc:contributor>Ferrando-May, Elisa</dc:contributor>
    <dcterms:rights rdf:resource="https://rightsstatements.org/page/InC/1.0/"/>
    <dc:creator>Kolmar, Harald</dc:creator>
    <dcterms:title>General mutagenesis/gene expression procedure for the construction of variant immunoglobulin domains in Escherichia coli : Production of the Bence-Jones protein REIv via fusion to beta-lactamase</dcterms:title>
    <dcterms:hasPart rdf:resource="https://kops.uni-konstanz.de/bitstream/123456789/22065/2/Darom_222146_MD%20in%20KOPS%20edit.pdf"/>
    <dc:contributor>Hennecke, Frank</dc:contributor>
    <bibo:uri rdf:resource="http://kops.uni-konstanz.de/handle/123456789/22065"/>
    <dc:contributor>Fritz, Hans-Joachim</dc:contributor>
    <dc:contributor>Kolmar, Harald</dc:contributor>
    <dc:creator>Ferrando-May, Elisa</dc:creator>
    <void:sparqlEndpoint rdf:resource="http://localhost/fuseki/dspace/sparql"/>
    <dc:rights>terms-of-use</dc:rights>
    <dc:creator>Wippler, Jürgen</dc:creator>
    <dc:date rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2013-03-20T10:49:04Z</dc:date>
    <dcterms:available rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2013-03-20T10:49:04Z</dcterms:available>
  </rdf:Description>
</rdf:RDF>

Interner Vermerk

xmlui.Submission.submit.DescribeStep.inputForms.label.kops_note_fromSubmitter

Kontakt
URL der Originalveröffentl.

Prüfdatum der URL

Prüfungsdatum der Dissertation

Finanzierungsart

Kommentar zur Publikation

Allianzlizenz
Corresponding Authors der Uni Konstanz vorhanden
Internationale Co-Autor:innen
Universitätsbibliographie
Nein
Begutachtet
Diese Publikation teilen