Molekularbiologische und biochemische Untersuchungen zur Aufnahme von osmoprotektiven Substanzen bei Corynebacterium glutamicum
Peter, H. (Corresponding author)
Publikationen vor 2000; PRE-2000; Retrocat
Jülich Forschungszentrum Jülich GmbH Zentralbibliothek, Verlag 1997
120 p.
Report
Book
Addenda
Berichte des Forschungszentrums Jülich 3460
OpenAccess
OpenAccess
Please use the identifier: http://hdl.handle.net/2128/22241 in citations.


The Gram-positive soil bacterium Corynebacterium glutamicum accumulates the osmoprotective substances glycine betaine, proline and ectoine under conditions of high osmolality. It was shown, that uptake of these solutes is mediated by four different secondary uptake systems. After isolation and sequencing of the corresponding genes, the kinetical function ofthe carrierproteins and their physiological relevance was studied. The transport system BetP mediates uptake of glycine betaine (K$_{m}$ = 8,6 $\mu$M) coupled to the cotransport of sodium (K$_{m}$ = 4,1 $\mu$M). Uptake activity of BetP is strongly regulated in dependence ofthe external osmolality. The regulation was studied by stepwise truncations of the C- and N-terminal domains. While N-terminal deletions led to a shift ofthe optimal activity into the range ofhigher osmolality, the C-terminal deletion of 12 amino acid residues revea1ed a shift ofthe optimal activity into the range of lower osmolality. Truncation of 23C-terminal amino acid residues led to a lowered affmity for the co substrate sodium and to the loss of the regulation. Additionally the affmity for glycine betaine was lowered, when 32 C-terminal amino acid residues were truncated. After deletion ofthe whole C-terminal domain no uptake activity could be observed anymore. The uptake system PutP catalyzes only transport of proline (K$_{m}$ = 7,6 $\mu$M) and is sodium dependend (K$_{m}$ = 1,3 $\mu$M). In contrast to BetP uptake acitivity of PutP is not osmotically regulated. ProP mediates proline (K$_{m}$ = 48 $\mu$M) and ectoine (K$_{m}$ = 132 $\mu$M) uptake coupled to the cotransport of protons and is osmotically regulated similar to BetP. The secondary structure prediction of ProP revealed the existence of hydrophilie C-and N-terminal domains as observed in the case of BetP. Additionally ProP is regulated on the level of expression, however no conditions of induction were found. EctP is a sodium coupled uptake system, which is constitutively expressed and osmotically regulated on the level of activity in the same manner as BetP and ProP. EctP shares a high degree of identity to BetP, but in contrast to the glycine betaine permease BetP it shows the highest affmity to ectoine (K$_{m}$ = 63 $\mu$M) followed by glycine betaine (K$_{m}$ = 333 $\mu$M) and proline (K$_{m}$ = 1200 $\mu$M). The physiological characterisation of different deletion strains of $\textit{C. glutamicum}$ showed, that the existence of the low affmity uptake system EctP is sufficient for osmoadaptation of the organism.