Substrate and phospholipid specificity of the purified mannitol permease of Escherichia coli.
D-Mannitol is transported and phosphorylated by a specific enzyme II of the phosphotransferase system of Escherichia coli. This protein was purified previously in detergent solution and has been partially characterized. As one approach in understanding the structure and mechanism of this enzyme/permease, we have tested a number of sugar alcohols and their derivatives as substrates and/or inhibitors of this protein. Our results show that the mannitol permease is highly, but not absolutely, specific for D-mannitol. Compounds accepted by the enzyme include those with substitutions in the C-2(= C-5) position of the carbon backbone of the natural substrate as well as D-mannonic acid, one heptitol and one pentitol. All of these compounds were both inhibitors and substrates for the mannitol permease except for D-mannoheptitol, which was an inhibitor but was not phosphorylated by the enzyme. No compound examined, however, exhibited an affinity for the enzyme as high as that for its natural substrate. We have also investigated the phospholipid requirements of the mannitol permease using phospholipids purified from E coli. The purified protein was significantly activated by phosphatidylethanolamine, but little activation was observed with phosphatidylglycerol or cardiolipin. These observations partially delineate requirements for interaction of sugar alcohols and phospholipids with the mannitol permease. They suggest approaches for the design of specific active site probes for the protein, and strategies for stabilizing the enzyme's activity in vitro.
D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D008353:Mannitol / Q000378:metabolism; D009004:Monosaccharide Transport Proteins; D010728:Phosphoenolpyruvate / Q000494:pharmacology; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism; D010766:Phosphorylation; D013379:Substrate Specificity
D029968:Escherichia coli Proteins; D009004:Monosaccharide Transport Proteins; D010743:Phospholipids; D008353:Mannitol; D010728:Phosphoenolpyruvate; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C022358:mannitol PTS permease, E coli