Febs Letters Volume 63, Number 1-2./Volume 64, Number 1-2. [antikvár]

Volume 63, number 1 l-EBS LETTERS March 1976 THE NATURE AND CONTROL OF CARBOHYDRATE UPTAKE BY ESCHERICHIA COLI* H. L KORNBERG Department of Biochemistry, University of Leicester UK** Received 24 December 1975 The uptake of carbohydrates from their growth media by /;. coii is brought about by two main processes. Both require the expenditure of metabolic energy; since the hydrophilic carbon compounds tliat serve as nutrients cannot traverse the hydrophobic barrier presented by the cell membrane unaided, both processes also necessitate the presence of specific proteins in that membrane. In the first of these processes, metabolic energy is transduced, in a manner not yet fully established, to effect the vectorial translocation of the carbohydrate but without changing its chemical identity. If the metabolism of that carbohydrate is prevented by the presence of some mutation that blocks an early step, the carbohydrate thus taken up 'actively' can be shown to accumulate, in concentration many times that present in the external miUeu. For example, the 'active' transport of labelled galactose by an E. coii mutant devoid of galactokinase activity was shown by Horecker, Thomas and Monod already some 15 years ago [I] to lead to a massive accumulation of the sugar in the cell; if unlabelled galactose was now added, the labelled material readily exchanged with it and the radioactivity was thus displaced from the cell. Similar experiments have been done with mutants impaired in the catabolism (but not the uptake) of arabinose. In the presence of an energy source, such mutants take up labelled arabinose and also concen- ¦ To avoid duplication, only 4 out of the 26 slides that accompanied the oral presentation are reproduced here; these 4 have not been published previously. References to the data contained in the remainder are given in the text. ** Present address: Department of Biochemistry, University of Cambridge, UK. trate it many-fold; again, the addition of unlabelled arabinose virtually totally 'washes out' the labelled sugar from the cell [2]. This shows that indeed nothing happens chemically to the sugar, and that it is taken up, unchanged, by the mutant. When the experiment is performed with starved bacteria, in the absence of an energy source, it is found that there is an uptake of H* ion concomitant with the uptake of the sugar; both accumulation of sugar and H^ uptake are arrested by the addition of agents that uncouple oxidative phosphorylation, such as CICCP [3]. Findings such as tliese very strongly support the view, first formulated by Dr Peter Mitchell [4], that the inward movement of the sugar is energised by a transmembrane gradient of protons and/or of electrical charge. Although there are a number of carbohydrates that are thus taken up by 'active transport', a second, and different, mechanism is also found in E.coli and in other bacteria capable of anaerobic growth. This also requires the input of metabolic energy, but the source of that energy is not an electro-chemical gradient but the metabolite phosphoenolpyruvate (PEP), which donates its pliosphate stoichometrically to the sugar. In consequence, the sugar taken up appears inside the cell not in the form in which it is present in the medium but as its phosphate ester; pyruvate is also formed as the other product of the reaction. The main features of this process, which was discovered by Dr Saul Roseman and his colleagues [S] and was designated the 'PEP-dependent phosphotransferase system' (abbreviated PT-system from now on) are illustrated in fig.l. The system includes at least five components although other constituents of the overall process are still being brought to light in the course of purifica- North-Holland Publishing Company - Amsterdam 1