salt-induced ABC transporter Ota of the methanogenic archaeon Methanosarcina mazei Gö1 is a glycine betaine transporter

• Published in: FEMS microbiology letters 2007-12, Vol.277 (1), p.44-49
• Main Authors: Schmidt, Silke, Pflüger, Katharina, Kögl, Stephanie, Spanheimer, Regina, Müller, Volker
• Format: Article
• Language: English
• Subjects: ABC transporter; ABC transporters; Archaea; Archaeal Proteins - genetics; Archaeal Proteins - metabolism; ATP-Binding Cassette Transporters - metabolism; betaine; Betaine - metabolism; Cloning, Molecular; Culture Media; E coli; Escherichia coli; Escherichia coli - genetics; Escherichia coli - growth & development; Escherichia coli - metabolism; Glycine; Glycine betaine; Kinetics; Membrane Transport Proteins - metabolism; methanogens; Methanosarcina - drug effects; Methanosarcina - genetics; Methanosarcina - metabolism; Methanosarcina mazei; Microbiology; osmoregulation; Ota; Proline; Protein transport; Sarcosine; Sodium Chloride - pharmacology; Sugar; Transport; Western blotting
• ISSN: 0378-1097; 1574-6968
• DOI: 10.1111/j.1574-6968.2007.00938.x

The genes encoding the three subunits of the primary ABC transporter Ota of the methanogenic archaeon Methanosarcina mazei Gö1 were cloned in an expression vector (pBAD24) and transformed into the glycine betaine transport-negative mutant Escherichia coli MKH13. Ota was produced as demonstrated by Western blotting. Uptake studies revealed that Ota catalyzed the transport of glycine betaine in E. coli MKH13(pBAD-Ota) with a Km of 10±5 μM and a maximal velocity of 1.5±0.5 nmol min⁻¹ mg protein⁻¹. Transport was ATP dependent. Ota was activated by salinity gradients, but only marginally by sugar gradients across the membrane. Glycine betaine transport was inhibited to a small extent by an excess of dimethylglycin or proline betaine, but not by sarcosine or glycine.