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Selenomethionine incorporation in proteins expressed in Lactococcus lactis.

ABSTRACT: Lactococcus lactis is a promising host for (membrane) protein overproduction. Here, we describe a protocol for incorporation of selenomethionine (SeMet) into proteins expressed in L. lactis. Incorporation efficiencies of SeMet in the membrane protein complex OpuA (an ABC transporter) and the soluble protein OppA, both from L. lactis, were monitored by mass spectrometry. Both proteins incorporated SeMet with high efficiencies (>90%), which greatly extends the usefulness of the expression host L. lactis for X-ray crystallography purposes. The crystal structure of ligand-free OppA was determined at 2.4 A resolution by a semiautomatic approach using selenium single-wavelength anomalous diffraction phasing.

SUBMITTER: Berntsson RP

PROVIDER: S-EPMC2771314 | biostudies-literature | 2009 May

REPOSITORIES: biostudies-literature

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Selenomethionine incorporation in proteins expressed in Lactococcus lactis.

Berntsson Ronnie P-A RP Alia Oktaviani Nur N Fusetti Fabrizia F Thunnissen Andy-Mark W H AM Poolman Bert B Slotboom Dirk-Jan DJ

Protein science : a publication of the Protein Society 20090501 5

Lactococcus lactis is a promising host for (membrane) protein overproduction. Here, we describe a protocol for incorporation of selenomethionine (SeMet) into proteins expressed in L. lactis. Incorporation efficiencies of SeMet in the membrane protein complex OpuA (an ABC transporter) and the soluble protein OppA, both from L. lactis, were monitored by mass spectrometry. Both proteins incorporated SeMet with high efficiencies (>90%), which greatly extends the usefulness of the expression host L. ...[more]

PMID: 19388077

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