Studies on the α‐Methylglucoside Permease of Escherichia coli

The sequential events which occur in the membrane of Escherichia coli and lead to the antecedent appearance of α‐methylglucoside 6‐phosphate when cells are exposed to extracellular α‐methylglucoside have been analyzed. It has been shown that phosphorylation is the rate limiting step in α‐methylglucoside phosphate synthesis in vivo . However, phosphorylation of α‐methylglucoside was not found to be the only step occurring in the thickness of membrane. It was shown that a step of transport was necessarily taking place before phosphorylation. In the absence of phosphorylation (due to genetic inactivation of any part of the enzymatic complex devoted to phosphorylation, or to chemical inactivation of one among its components), this step was expressed as a facilitated diffusion of unaltered α‐methylglucoside. The demonstration of this facilitated passage rests on the following evidence: equalization of α‐methylglucoside concentrations in intracellular and extracellular compartments faster than when due to free passive diffusion alone; requirement of the interaction of α‐methylglucoside with a stereospecific site, distinct from E II , with a K m of 20 μM; finally, existence of counterflows suggesting that the passage was ensured by a mobile carrier, mediating both influx and efflux of α‐methylglucoside. This step of stransport was shown to be required in intact cells for a normal efficiency of phosphorylation in vivo . However it was not expressed in vivo as a facilitated diffusion but very likely as a facilitated access to the site of phosphorylation of E II . From these findings, and from an analysis of various mutants, it was concluded that a concerted change of conformation occurs when E II was phosphorylating, which renders the step of transport unlimiting. Some consequences of the dissociation of the α‐methylglucoside phosphate synthesis into two sequential steps are discussed. Mainly this model provides a simple explanation for the polarity exhibited by E II , the absence of phosphorylation of intracellular α‐methylglucoside and its continuous exchange.