Immediate inhibition of sucrose uptake inCorynebacterium glutamicumin response to intracellular glucose-6-phosphate accumulation requires theptsGencoded EII-permease

Abstract

Running title: Coordination of PTS sugar uptake and central metabolism in C. glutamicum 22 Content category: Biochemistry and Physiology 23Abbreviations: cdm -cell dry mass; WT, wild-type. 24 ABSTRACT 28Corynebacterium glutamicum co-metabolizes most carbon sources, such as glucose and 29 sucrose. Uptake of those sugars by the PTS involves a glucose-and a sucrose-specific 30 permease EII Glc (ptsG) and EII Suc (ptsS), respectively. Block of glycolysis by deletion of pgi 31 (encodes phosphoglucoisomerase) redirects glucose-driven carbon flux towards pentose 32 phosphate pathway. C. glutamicum Δpgi grows poorly with glucose but has unaffected, good 33 growth with sucrose. However, addition of glucose to sucrose-cultivated C. glutamicum Δpgi 34 immediately arrested growth via inhibition of the EII Suc -mediated sucrose uptake and 35 reduction of ptsS-mRNA amounts. Kinetic analyses revealed that sucrose uptake inhibition in 36 C. glutamicum Δpgi took place within 15 s after glucose addition. We show that inhibition of 37 PTS-mediated sucrose uptake occurs as direct response to glucose-6-P accumulation. 38Moreover, addition of non-PTS substrates, which are metabolized to glucose-6-P such as 39 maltose or glucose-6-P itself (uptake was enabled by heterologously produced UhpT), led to 40 similar growth and sucrose uptake inhibition as glucose addition. Despite EII Glc not being 41 involved in uptake of these substrates, negative effects on sucrose uptake after addition of 42 maltose and glucose-6-P were absent in the EII Glc -deficient strain C. glutamicum ΔpgiΔptsG. 43These results show that the ptsG-encoded EII Glc is part of a novel mechanism for perception of 44 intracellular glucose-6-P accumulation and instantaneous inhibition of EII Suc -mediated 45 sucrose uptake in C. glutamicum. This novel mode of control of PTS activity by an early 46 glycolytic metabolite probably allows efficient adaptation of sugar uptake to the capacity of 47 IMPORTANCE 50Coordination of substrate uptake and metabolism are a prerequisite for efficient co-utilization 51 of substrates, a trait typical for the Gram-positive C. glutamicum. Sucrose uptake via the PTS 52 permease EII Suc in this organism immediately was inhibited in response to intracellular 53 accumulation of the glycolysis intermediate glucose-6-phosphate. This inhibition depends 54 exclusively on the presence but not activity of the PTS permease EII Gluc . Thus, C. glutamicum 55 possesses a novel, immediate, and PTS-dependent way to control and coordinate both uptake 56 and metabolization of multiple substrates by monitoring of their metabolic levels in the cell. 57This offers new insights and interesting concepts for a further rational engineering of this 58 industrially important production organism and exemplifies a putative general strategy of 59 bacteria for the coordination of sugar uptake and central metabolism. 60 61 sucrose metabolism. As characterized here, sucrose in this bacterium is exclusively taken up 126 6 via the sucrose-specific, ptsS-encoded EII Suc with a ...