Comparison of the functional properties of trimeric and monomeric CaiT of Escherichia coli

Bracher, Susanne; Hilger, Daniel; Guerin, Kamila; Polyhach, Yevhen; Jeschke, Gunnar; Krafczyk, Ralph; Giacomelli, Giacomo und Jung, Heinrich (2019): Comparison of the functional properties of trimeric and monomeric CaiT of Escherichia coli. In: Scientific Reports, Bd. 9, 3787

Abstract

Secondary transporters exist as monomers, dimers or higher state oligomers. The significance of the oligomeric state is only partially understood. Here, the significance of the trimeric state of the L-carnitine/gamma-butyrobetaine antiporter CaiT of Escherichia coli was investigated. Amino acids important for trimer stability were identified and experimentally verified. Among others, CaiT-D288A and -D288R proved to be mostly monomeric in detergent solution and after reconstitution into proteoliposomes, as shown by blue native gel electrophoresis, gel filtration, and determination of intermolecular distances. CaiT-D288A was fully functional with kinetic parameters similar to the trimeric wild-type. Significant differences in amount and stability in the cell membrane between monomeric and trimeric CaiT were not observed. Contrary to trimeric CaiT, addition of substrate had no or only a minor effect on the tryptophan fluorescence of monomeric CaiT. The results suggest that physical contacts between protomers are important for the substrate-induced changes in protein fluorescence and the underlying conformational alterations.

• Dokumententyp: Zeitschriftenartikel
• Fakultät: Biologie > Department Biologie I
• Themengebiete: 500 Naturwissenschaften und Mathematik > 570 Biowissenschaften; Biologie
• ISSN: 2045-2322
• Sprache: Englisch