Selenomethionine incorporation in proteins expressed in Lactococcus lactis

✍ Scribed by Ronnie P.-A. Berntsson; Nur Alia Oktaviani; Fabrizia Fusetti; Andy-Mark W. H. Thunnissen; Bert Poolman; Dirk-Jan Slotboom

• Book ID: 105356787
• Publisher: Cold Spring Harbor Laboratory Press
• Year: 2009
• Tongue: English
• Weight: 589 KB
• Volume: 18
• Category: Article
• ISSN: 0961-8368
• DOI: 10.1002/pro.97

✦ Synopsis

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 Å resolution by a semiautomatic approach using selenium single‐wavelength anomalous diffraction phasing.