Uptake of glutamate in <i>Corynebacterium glutamicum</i>

Krämer, Reinhard; Lambert, Camille

doi:10.1111/j.1432-1033.1990.tb19489.x

Cited by 56 publications

(32 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, low levels of DCCD increased the membrane potential, presumably because protons that were pumped out by the respiratory chain could not flow into the cells through the Fo proton channel. Potential values of up to 210 mV were reached, as in previous studies (13). The export of arsenite was inhibited at very low ATP concentrations (Fig.…”

Section: Discussionsupporting

confidence: 53%

Arsenic efflux governed by the arsenic resistance determinant of Staphylococcus aureus plasmid pI258

Bröer

et al. 1993

J Bacteriol

View full text Add to dashboard Cite

The arsenic resistance operon of Staphylococcus aureus plasmid p1258 determined lowered net cellular uptake of "As by an active efflux mechanism. Arsenite was exported from the cells; intracellular arsenate was first reduced to arsenite and then transported out of the cells. (15) showed that efflux determined by the E. coli plasmid R773 arsenic resistance operon was ATP dependent.Arsenic resistance determinants were cloned and sequenced. The E. coli plasmid R773 ars operon consists of five genes (arsR, arsD, arsA, arsB, and arsC) (3,18,22,26,29), but only three genes (arsR, arsB, and arsC) were found on Staphylococcus plasmids (9,20). The arsR, arsB, and arsC gene products of both operons have similarities in sequence and function. The arsR gene encodes a repressor protein in both organisms (20,28). The R773 (E. coli) and p1258 (S. aureus) ArsR proteins have 30% identical amino acid residues. The E. coli ArsB protein is a membrane protein with 12 transmembrane a-helices (18,23,30) and has amino acid residues 58% identical to those of the staphylococcal ArsB protein (9). The arsC gene encodes an arsenateto-arsenite reductase (10) in both microorganisms, although the sequence similarity is slight (with only 18% identical residues between the S. aureus and E. coli versions). The function of the arsenate reductase explains why arsC is needed only for arsenate resistance (3, 9, 26). ArsC reduces arsenate to arsenite, which subsequently is transported out of the cells (10). The arsenate oxyanion itself is not transported out of the cell (10). The E. coli R773 ars operon includes the extra gene arsA, which determines an ATPase subunit (3,8). ArsA has been purified and functions as an arsenite-or antimonite-stimulated soluble ATPase (19),

show abstract

Section: Discussionsupporting

confidence: 53%

Arsenic efflux governed by the arsenic resistance determinant of Staphylococcus aureus plasmid pI258

Bröer

et al. 1993

J Bacteriol

View full text Add to dashboard Cite

show abstract

“…(d) The ion dependence with respect to stimulation by Na' or K + was absolutely identical (not shown). (e) Glutamate uptake in both cases was coupled to the same energy source as will be shown in the accompanying paper [22].…”

Section: Discussionmentioning

confidence: 99%

Uptake of glutamate in Corynebacterium glutamicum

Krämer

Lambert

Hoischen

et al. 1990

European Journal of Biochemistry

Self Cite

View full text Add to dashboard Cite

The active uptake system for glutamate in Corynebacterium glutamicum is inducible by growth on glutamate as sole energy and carbon source and is also susceptible to catabolite repression by glucose. The basic level of uptake activity is low in glucose‐grown cells (1.5 nmol · mg dry mass−1· min−1), it is intermediate when acetate is the carbon source (3.8 nmol · mg dry mass−1· min−1) and becomes fully induced by glutamate (15 nmol · mg dry mass−1· min−1). In all cases the uptake has, except for different Vmax values, identical kinetic and energetic properties, and is characterized by a low apparent Km value of 0.5–1.3 μM and by high substrate specificity. The transported substrate species is the deprotonated form which can also be concluded from the extremely high pH optimum of transport above pH 9. Glutamate uptake in cells grown in media with low K+ concentration is not influenced by external Na+ but is drastically stimulated by addition of K+. Stimulation by K+ could be separated into two different mechanisms. (a) Addition of K+ increases the internal pH, thereby stimulating glutamate uptake which is regulated by the internal pH in C. glutamicum. The apparent pK of the internal ‘pH switch’ is 6.6; below this value, uptake of glutamate is inhibited. (b) Internal K+ also directly promotes glutamate uptake. Effective uptake of glutamate can be observed only when the cytosolic K+ concentration exceeds a threshold value of about 200 mM. Stimulation of glutamate uptake by external K+ is not due to functional coupling of K+ and glutamate transport but reveals the necessity to replenish the internal K+ pool.

show abstract

“…The glutamate uptake system of C. glutamicum is clearly different from the glutamine uptake system of this organism (14). Moreover, growth with glutamate as a carbon source is directly dependent on the capacity of the glutamate transport system (21). Therefore, mutagenized cells of the wild type of C. glutamicum were replica plated on minimal medium with either glutamate or glutamine as the carbon source.…”

Section: Resultsmentioning

confidence: 99%

“…Functional studies with intact cells on the kinetics and energetics of several amino acid carriers of C. glutamicum, including that of the glutamate uptake system, have been carried out (for a review, see reference 20). This uptake system (i) exhibits high affinity, (ii) results in extremely high internal glutamate accumulation, (iii) is irreversible, and (iv) has activity directly correlated to the cytosolic ATP content of the cell (21). These results were interpreted in terms of a primary, binding proteindependent, ATP-driven uptake system for glutamate.…”

mentioning

confidence: 97%

Structure of the gluABCD cluster encoding the glutamate uptake system of Corynebacterium glutamicum

et al. 1995

View full text Add to dashboard Cite

To assess the mechanism and function of the glutamate uptake system of gram-positive Corynebacterium glutamicum, a mutant deficient in glutamate uptake was isolated and was then used to isolate a DNA fragment restoring this deficiency. In a low-copy-number vector, this fragment resulted in an increased glutamate uptake rate of 4.9 nmol/min/mg (wild type, 1.5 nmol/min/mg). In addition, carbon source-dependent regulation of the glutamate uptake system was determined with the fragment, showing that the entire structures required for expression and control reside on the fragment isolated. Sequencing of 3,977 bp revealed the presence of a four-gene cluster (gluABCD) with deduced polypeptide sequences characteristic of a nucleotide-binding protein (GluA), a periplasmic binding protein (GluB), and integral membrane proteins (GluC and GluD), identifying the glutamate transporter as a binding protein-dependent system (ABC transporter). This identification was confirmed by the kinetic characteristics obtained for cells grown in the presence of globomycin, which exhibited an increased K m of 1,400 M (without globomycin, the K m was 1.5 M) but a nearly unaltered maximum velocity. By applying gene-directed mutagenesis, a strain with the entire cluster deleted was constructed. With this mutant, the glutamate uptake rate was reduced from 1.4 to less than 0.1 nmol/min/mg, which is proof that this system is the only relevant one for glutamate uptake. With this strain, the glutamate excretion rate was unaffected (18 nmol/min/mg), showing that no component of gluABCD is involved in export but rather that a specific machinery functions for the latter purpose.Gram-positive Corynebacterium glutamicum is in use for the large-scale production of amino acids, in particular for glutamate. However, compared with what is known about many other bacteria, molecular details on the amino acid transport systems of this bacterium are severely restricted. So far, only the structure of the lysine uptake carrier, lysI, is known (33). Functional studies with intact cells on the kinetics and energetics of several amino acid carriers of C. glutamicum, including that of the glutamate uptake system, have been carried out (for a review, see reference 20). This uptake system (i) exhibits high affinity, (ii) results in extremely high internal glutamate accumulation, (iii) is irreversible, and (iv) has activity directly correlated to the cytosolic ATP content of the cell (21). These results were interpreted in terms of a primary, binding proteindependent, ATP-driven uptake system for glutamate. Interestingly, the total catalytic activity of the system is induced when cells are grown on glutamate but it is subject to catabolite repression by glucose (22).Binding protein-dependent systems belong to the large family of ABC transporters (for a review, see reference 11). These multicomponent systems typically consist of two membraneinserted subunits (domains), one component inside the cytoplasm which carries the ATP-binding site, and finally the binding protein ...

show abstract

Uptake of glutamate in Corynebacterium glutamicum

Cited by 56 publications

References 34 publications

Arsenic efflux governed by the arsenic resistance determinant of Staphylococcus aureus plasmid pI258

Arsenic efflux governed by the arsenic resistance determinant of Staphylococcus aureus plasmid pI258

Uptake of glutamate in Corynebacterium glutamicum

Structure of the gluABCD cluster encoding the glutamate uptake system of Corynebacterium glutamicum

Contact Info

Product

Resources

About