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A Schlösser

Publications and source records attributed to A Schlösser.

15 recordsLinked to original sources
PubMed2002

The novel Streptomyces olivaceoviridis ABC transporter Ngc mediates uptake of N-acetylglucosamine and N,N'-diacetylchitobiose.

During cultivation in the presence of N-acetylglucosamine or chitin, Streptomyces olivaceoviridis mycelium efficiently takes up [(14)C]-labelled N-acetylglucosamine. Uptake of the labelled compound can be completely inhibited by unlabelled N-acetylglucosamine and partially by chitobiose. After extraction of the membrane with Triton X-100, two forms of a protein that binds to N-acetylglucosamine and N, N'-diacetylchitobiose (chitobiose) were purified to homogeneity by two consecutive rounds of anionic exchange chromatography. The protein was named NgcE. Using surface plasmon resonance, its binding parameters were determined. It showed highest affinity for N-acetylglucosamine (K(D)=8.28 x 10(-9) M) and for chitobiose (K(D)=2.87 x 10(-8) M). Varying equilibrium dissociation constants in the micromolecular range were ascertained for chitotetraose (K(D)=4.5 x 10(-6) M), chitopentaose (K(D)=1.03 x 10(-6) M) and chitohexaose (K(D)=3.02 x 10(-6) M); the lowest value was measured for chitotriose (K(D)=19.4 x 10(-6) M). After having determined the sequences of several internal peptides from the binding protein by Edman degradation, the corresponding ngcE gene, which encodes a predicted lipid-anchored protein, was identified by reverse genetics. Using a genomic phage library of S. olivaceoviridis genes encoding two other membrane proteins (named NgcF and NgcG) were identified adjacent to ngcE. Each of these is predicted to have six membrane-spanning helices and a consensus motif for integral membrane proteins characteristic of ABC transporters. In addition, the gene for a predicted regulator protein (NgcR) was detected. The ngcEFG operon lacks a gene for an ATP-hydrolysing protein. NgcE is a new member of the CUT-1 family of ABC transporters for carbohydrates. Comparative studies of the wild-type and a mutant strain carrying an insertion within the ngc operon clearly demonstrate that the Ngc system mediates the uptake of N-acetylglucosamine and chitobiose in vivo.

ATP-Binding Cassette Transporters↗
PubMed2001

Synthesis of the Streptomyces lividans maltodextrin ABC transporter depends on the presence of the regulator MalR.

During growth with maltotriose or amylose, Streptomyces lividans and Streptomyces coelicolor A3(2) synthesize a maltodextrin uptake system with highest specificity for maltotriose. The transport activity is absent in mutants of S. coelicolor A3(2) lacking a functional MalE binding protein. Cloning and sequencing data suggest that the mal operon of S. coelicolor A3(2) corresponds to the one of S. lividans and that the deduced S. lividans Reg1 amino acid sequence is identical to that of MalR from S. coelicolor A3(2). It can be concluded that both strains have the same ABC transport system for maltodextrins. The S. lividans malR was cloned in Escherichia coli in frame with six histidine-encoding codons. The resulting, purified 6HisMalR(SI) was shown to bind to two motifs within the S. lividans malR-malE intergenic region and to dissociate in the presence of maltopentaose.

ATP-Binding Cassette Transporters↗
PubMed2000

Binding characteristics of CebR, the regulator of the ceb operon required for cellobiose/cellotriose uptake in Streptomyces reticuli.

The Streptomyces reticuli Avicelase (cellulase, Cell) hydrolyzes crystalline cellulose to cellooligomers, cellobiose and cellotriose which are taken up by mycelia via an ABC transport system (Ceb) induced during growth with cellobiose or cellulose. The cebR gene located upstream of the cebEFG operon was cloned in Escherichia coli in frame with six histidine-encoding codons. The resulting purified fusion protein was shown to bind to a motif of 23 bp, including a perfect 18-bp palindrome situated upstream of the cebEFG. Cytoplasmic extracts of induced, but not of uninduced S. reticuli protected the same DNA motif. Release of the CebR regulator from its operator occurs upon addition of cellopentaose which can be assumed to act as inducer within the mycelia.

ATP-Binding Cassette Transporters↗
PubMed2000

MsiK-dependent trehalose uptake in Streptomyces reticuli.

During cultivation in the presence of trehalose Streptomyces reticuli expresses an inducible, highly specific trehalose uptake system that is absent in Streptomyces lividans. A palmitated trehalose-binding protein was identified in the cytoplasmic membrane of mycelia, extracted with the detergent Triton X-100 and purified using a trehalose affinity matrix. Immunological studies showed that within S. reticuli the synthesis of the ATP-binding protein MsiK is induced by trehalose. The data suggest that MsiK assists the trehalose ABC transporter, like the previously described ABC transport systems for maltose and cellobiose/cellotriose, respectively.

ATP-Binding Cassette Transporters↗
PubMed1999

Characterization of the binding protein-dependent cellobiose and cellotriose transport system of the cellulose degrader Streptomyces reticuli.

Streptomyces reticuli has an inducible ATP-dependent uptake system specific for cellobiose and cellotriose. By reversed genetics a gene cluster encoding components of a binding protein-dependent cellobiose and cellotriose ABC transporter was cloned and sequenced. The deduced gene products comprise a regulatory protein (CebR), a cellobiose binding lipoprotein (CebE), two integral membrane proteins (CebF and CebG), and the NH2-terminal part of an intracellular beta-glucosidase (BglC). The gene for the ATP binding protein MsiK is not linked to the ceb operon. We have shown earlier that MsiK is part of two different ABC transport systems, one for maltose and one for cellobiose and cellotriose, in S. reticuli and Streptomyces lividans. Transcription of polycistronic cebEFG and bglC mRNAs is induced by cellobiose, whereas the cebR gene is transcribed independently. Immunological experiments showed that CebE is synthesized during growth with cellobiose and that MsiK is produced in the presence of several sugars at high or moderate levels. The described ABC transporter is the first one of its kind and is the only specific cellobiose/cellotriose uptake system of S. reticuli, since insertional inactivation of the cebE gene prevents high-affinity uptake of cellobiose.

ATP-Binding Cassette Transporters↗
PubMed1998

The ABC maltose transporter.

Bacterial ATP-binding cassette (ABC) transporters and their homologues in eukaryotic cells form one of the largest superfamilies known today. They function as primary pumps that couple substrate translocation across the cytoplasmic membrane to ATP hydrolysis. Although ABC transporters have been studied for more than three decades, the structure of these multi-component systems is unknown, and the mechanism of transport is not understood. This article reviews one of the most widely studied ABC systems, the maltose transporter of Escherichia coli. A first structural model of the transport channel allows discussion of possible mechanisms of transport. In addition, recent experimental evidence suggests that regulation of gene expression and transport activity is far more complex than expected.

ATP-Binding Cassette Transporters↗
PubMed1997

The Streptomyces ATP-binding component MsiK assists in cellobiose and maltose transport.

Streptomyces reticuli harbors an msiK gene which encodes a protein with an amino acid identify of 90% to a corresponding protein previously identified in Streptomyces lividans. Immunological studies revealed that S. lividans and S. reticuli synthesize their highest levels of MsiK during growth with cellobiose, but not with glucose. Moreover, moderate amounts of MsiK are produced by both species in the course of growth with maltose, melibiose, and xylose and by S. lividans in the presence of xylobiose and raffinose. In contrast, a recently identified cellobiose-binding protein and its distantly related homolog were only found if S. reticuli or S. lividans, respectively, was cultivated with cellobiose. Uptake of cellobiose and maltose was tested and ascertained for S. reticuli and S. lividans, but not for an msiK S. lividans mutant. However, transformants of this mutant carrying the S. reticuli or S. lividans msiK gene on a multicopy plasmid had regained the ability to transport both sugars. The data show that MsiK assists two ABC transport systems.

ATP-Binding Cassette Transporters↗
PubMed1996

A lipid-anchored binding protein is a component of an ATP-dependent cellobiose/cellotriose-transport system from the cellulose degrader Streptomyces reticuli.

During cultivation in the presence of cellobiose or crystalline cellulose, Streptomyces reticuli expresses an inducible uptake system that transports cellobiose (K(m), 4 microM), cellotriose and, to a lesser degree, cellotetraose and cellopentaose. Cellobiose uptake is dependent on ATP and inhibited by N-ethylmaleimide. A binding protein was identified in its palmitylated form in the cytoplasmic membrane of mycelia. It could be extracted with the detergent Triton X-100 and purified by two subsequent anion-exchange chromatographies. It showed highest affinity (Kd, 1.5 microM) for cellobiose and cellotriose. The data suggest that cellobiose/cellotriose uptake is mediated by a membrane-anchored lipoprotein as a component of an ATP-binding-cassette-transporter system.

Adenosine Triphosphate↗
PubMed1995

TrkH and its homolog, TrkG, determine the specificity and kinetics of cation transport by the Trk system of Escherichia coli.

The corrected sequence of the trkH gene of Escherichia coli predicts that the TrkH protein is a hydrophobic membrane protein of 483 amino acid residues, of which 41% are identical to those of the homologous and functionally analogous TrkG protein. These two proteins form the transmembrane component of the Trk system for the uptake of K+. Each protein alone is sufficient for high-level Trk activity. When Trk is assembled with the TrkG protein, Rb+ and K+ are transported with a Km near or below 1 mM; however, the Vmax for Rb+ is only about 7% of that for K+. When Trk is formed with TrkH, the affinities for both for K+ and Rb+ are somewhat lower, and the Vmax for Rb+ is only 1% of that for K+ transport. The kinetics of transport in strains with wild-type alleles at trkG and at trkH suggest that both products participate in transport.

Amino Acid Sequence↗
PubMed1993

NAD+ binding to the Escherichia coli K(+)-uptake protein TrkA and sequence similarity between TrkA and domains of a family of dehydrogenases suggest a role for NAD+ in bacterial transport.

The nucleotide sequence of trkA, a gene encoding a surface component of the constitutive K(+)-uptake systems TrkG and TrkH from Escherichia coli, was determined. The structure of the TrkA protein deduced from the nucleotide sequence accords with the view that TrkA is peripherally bound to the inner side of the cytoplasmic membrane. Analysis by a dot matrix revealed that TrkA is composed of similar halves. The N-terminal part of each TrkA half (residues 1-130 and 234-355, respectively) is similar to the complete NAD(+)-binding domain of NAD(+)-dependent dehydrogenases. The C-terminal part of each TrkA half (residues 131-233 and 357-458, respectively) aligns with the first 100 residues of the catalytic domain of glyceraldehyde-3-phosphate dehydrogenase. Strong u.v. illumination at 252 nm led to cross-linking of NAD+ or NADH, but not of ATP to the isolated TrkA protein.

ATP-Binding Cassette Transporters↗
PubMed1991

Subcloning, nucleotide sequence, and expression of trkG, a gene that encodes an integral membrane protein involved in potassium uptake via the Trk system of Escherichia coli.

The trkG gene encodes a component of the K+ uptake system Trk and is located at 30.5 min inside the lambdoid prophage region rac of the Escherichia coli chromosome. trkG was subcloned, its nucleotide sequence was determined, and its product was identified in a minicell system. The open reading frame of 1,455 bp encodes a hydrophobic membrane protein with a calculated molecular weight of 53,493 that is predicted to contain up to 12 transmembrane helices. The trkG gene product behaved as a hydrophobic membrane protein; it was found exclusively in the membrane fraction of the minicells and its migration in sodium dodecyl sulfate-polyacrylamide gel electrophoresis was anomalous, indicating an apparent molecular weight of 35,000. The trkG gene contains an exceptionally high proportion of infrequently used codons, raising the question of the origin of this gene. trkG does not appear to be a prophage gene since no similarity was observed between the nucleotide sequence of trkG or the amino acid sequence of its product and the sequences of genes or proteins from bacteriophage lambda.

Amino Acid Sequence↗
PubMed1989

Specific cesium transport via the Escherichia coli Kup (TrkD) K+ uptake system.

Escherichia coli cells which contain a functional Kup (formerly TrkD) system took up Cs+ with a moderate rate and affinity. Kup is a separate K+ uptake system with relatively little discrimination in the transport of the cations K+, Rb+, and Cs+. Regardless of the presence or absence of Kup, K+-replete cells took up Cs+ primarily by a very low affinity mode, proportional to the ratio of the Cs+ and K+ concentrations in the medium.

Binding, Competitive↗
PubMedDate not supplied

Biochemical identification of a lipoprotein with maltose-binding activity in the thermoacidophilic Gram-positive bacterium Alicyclobacillus acidocaldarius.

Growth of the thermoacidophilic Gram-positive bacterium Alicyclobacillus acidocaldarius strain ATCC 27009 on maltose resulted in the increased production of a protein with apparent molecular mass of 40 kDa. By metabolic labelling with 14C-palmitic acid, the 40-kDa protein was identified as a lipoprotein. The protein exhibited maltose-binding activity at pH 3.5, as demonstrated by chromatography on cross-linked amylose. Partial amino acid sequence analysis revealed that the 40-kDa protein corresponds to the product of an open reading frame downstream from the amylase gene (amy) that displays similarity to enterobacterial maltose-binding proteins.

Amino Acid Sequence↗