A n t o n i e van L e e u w e n h o e k 4 5 ( 1 9 7 9 )
315
Glucose uptake by Klebsiella aerogenes The role of the glucose-phosphoenolpyruvate phosphotransferase transport system
R. W . O'BRIEN, O. M . NEIJSSEL AND D . W . TEMPEST Laboratorium voor Microbiologie, Universiteit van Amsterdam, Plantage Muidergracht 14, 1018 TV Amsterdam, The Netherlands The relationship between the uptake of glucose and the activity of the glucosephosphoenolpyruvate phosphotransferase in K. aerogenes NCTC 418 was investigated. At tow dilution rates in glucose-limited chemostat cultures the activity of the phosphotransferase was high, but decreased markedly with increasing growth rate. At dilution rates above 0.7 h - 1 glucose was transported at a rate greater than that which could be accounted for by the phosphotransferase activity. Addition of glucose (10 mM) to glucose-limited cultures showed that cells could transport glucose at a fast rate, irrespective of the growth rate. Further, glucose-sufficient cultures (either phosphate-limited or potassium-limited) transported glucose at rates 2 to 5 times greater than the activity of the phosphotransferase system. These results indicated that systems, other than the phosphotransferase, were available to K. aerogenes for the transport of glucose. Evidence for this was shown by lactose-limited cultures which possessed a relatively low activity of phosphotransferase, but were able to transport glucose at a fast rate. When such cultures were pulsed with an equimolar mixture of glucose and galactose they transported the glucose at a rate equivalent to the phosphotransferase activity; galactose was also transported at a rate equal to the difference in the rate of glucose uptake between lactose-limited cultures pulsed with glucose alone and with glucose plus galactose. Thus glucose appeared to be transported on the galactose permease. The above results suggest that the "high-affinity" phosphotransferase uptake system is responsible for glucose uptake when the extracellular concentration of glucose is low and that at high concentrations of glucose other transport systems, with a low affinity for glucose, play a significant role. A similar behaviour was observed with K. aerogenes strain NCIB 418 (supposedly identical to NCTC 418), NCIB 8805 and NCIB 8258. R. W. O'Brien thanks Z.W.O. for financial support.
Cell w a l l m e t a b o l i s m o f B a c i l l u s s u b t i l i s
W . R . DE BOER, F. J. KRUYSSEN AND J. T. M . WOUTERS Laboratorium voor Microbiologie, Universiteit van Amsterdam Plantage Muidergraeht 14, Amsterdam, The Netherlands Unlike earlier views favouring a rigid structure of the bacterial cell wall embracing the cytoplasmic membrane as an inflexible and inert box, it is now generally assumed that its main component, the peptidoglycan is subject to considerable metabolic activity, also after linkage to the pre-existing wall, resulting in a continuous reorientation and sliding of the wall polymers in the expanding surface of the growing cell.
316
A n t o n i e van L e e u w e n h o e k 45
(1979)
Recent data point to an accurate regulation of these processes in such a way as to maintain the order of the wall material throughout the wall outward from the cytoplasmic membrane. In growing cells, newly made peptidoglycan has been shown to be spread over the entire surface of the cell and its progeny in a strictly ordered manner (Pooley, 1976). Since new chains are laid down on the inside of the wall, an outward arrangement of consecutive layers of increasing age will result. In Bacillus subtilis and several other bacterial species the order between older and newer chains is lost in the exterior layers of the wall and r a n d o m excision by lytic enzymes does take place leading to turnover of wall material. The regulation and function of the process are poorly understood; the necessity for wall turnover related to growth and division does not seem to be obvious. In order to gain further insight into the mechanisms governing the regular growth of the bacterial wall0 we have analysed the kinetics of the loss of radioactivity from cells labeled specifically in their walls, after their transfer to nonradioactive medium.
B~ Time:
~
t
315
Glucose uptake by Klebsiella aerogenes The role of the glucose-phosphoenolpyruvate phosphotransferase transport system
R. W . O'BRIEN, O. M . NEIJSSEL AND D . W . TEMPEST Laboratorium voor Microbiologie, Universiteit van Amsterdam, Plantage Muidergracht 14, 1018 TV Amsterdam, The Netherlands The relationship between the uptake of glucose and the activity of the glucosephosphoenolpyruvate phosphotransferase in K. aerogenes NCTC 418 was investigated. At tow dilution rates in glucose-limited chemostat cultures the activity of the phosphotransferase was high, but decreased markedly with increasing growth rate. At dilution rates above 0.7 h - 1 glucose was transported at a rate greater than that which could be accounted for by the phosphotransferase activity. Addition of glucose (10 mM) to glucose-limited cultures showed that cells could transport glucose at a fast rate, irrespective of the growth rate. Further, glucose-sufficient cultures (either phosphate-limited or potassium-limited) transported glucose at rates 2 to 5 times greater than the activity of the phosphotransferase system. These results indicated that systems, other than the phosphotransferase, were available to K. aerogenes for the transport of glucose. Evidence for this was shown by lactose-limited cultures which possessed a relatively low activity of phosphotransferase, but were able to transport glucose at a fast rate. When such cultures were pulsed with an equimolar mixture of glucose and galactose they transported the glucose at a rate equivalent to the phosphotransferase activity; galactose was also transported at a rate equal to the difference in the rate of glucose uptake between lactose-limited cultures pulsed with glucose alone and with glucose plus galactose. Thus glucose appeared to be transported on the galactose permease. The above results suggest that the "high-affinity" phosphotransferase uptake system is responsible for glucose uptake when the extracellular concentration of glucose is low and that at high concentrations of glucose other transport systems, with a low affinity for glucose, play a significant role. A similar behaviour was observed with K. aerogenes strain NCIB 418 (supposedly identical to NCTC 418), NCIB 8805 and NCIB 8258. R. W. O'Brien thanks Z.W.O. for financial support.
Cell w a l l m e t a b o l i s m o f B a c i l l u s s u b t i l i s
W . R . DE BOER, F. J. KRUYSSEN AND J. T. M . WOUTERS Laboratorium voor Microbiologie, Universiteit van Amsterdam Plantage Muidergraeht 14, Amsterdam, The Netherlands Unlike earlier views favouring a rigid structure of the bacterial cell wall embracing the cytoplasmic membrane as an inflexible and inert box, it is now generally assumed that its main component, the peptidoglycan is subject to considerable metabolic activity, also after linkage to the pre-existing wall, resulting in a continuous reorientation and sliding of the wall polymers in the expanding surface of the growing cell.
316
A n t o n i e van L e e u w e n h o e k 45
(1979)
Recent data point to an accurate regulation of these processes in such a way as to maintain the order of the wall material throughout the wall outward from the cytoplasmic membrane. In growing cells, newly made peptidoglycan has been shown to be spread over the entire surface of the cell and its progeny in a strictly ordered manner (Pooley, 1976). Since new chains are laid down on the inside of the wall, an outward arrangement of consecutive layers of increasing age will result. In Bacillus subtilis and several other bacterial species the order between older and newer chains is lost in the exterior layers of the wall and r a n d o m excision by lytic enzymes does take place leading to turnover of wall material. The regulation and function of the process are poorly understood; the necessity for wall turnover related to growth and division does not seem to be obvious. In order to gain further insight into the mechanisms governing the regular growth of the bacterial wall0 we have analysed the kinetics of the loss of radioactivity from cells labeled specifically in their walls, after their transfer to nonradioactive medium.
B~ Time:
~
t
