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At least 19 recordsLinked to original sources

PubMed1984

Reconstitution of maltose chemotaxis in Escherichia coli by addition of maltose-binding protein to calcium-treated cells of maltose regulon mutants.

Maltose chemotaxis was reconstituted in delta malE cells lacking maltose-binding protein (MBP). Purified MBP was introduced into intact cells during incubation with 250 mM CaCl2 in Tris-hydrochloride buffer at 0 degrees C. After removal of extracellular CaCl2 and MBP, chemotaxis was measured with tethered bacteria in a flow chamber or with free-swimming cells in a capillary assay. About 20% of tethered cells responded to 10(-4) M maltose; the mean response times were about half those of CaCl2-treated wild-type cells (100 s as opposed to 190 s). In capillary tests, the maltose response of reconstituted cells was between 15 and 40% of the aspartate response, about the same percentage as in wild-type cells. The best reconstitution was seen with 0.5 to 1 mM MBP in the reconstitution mixture, which is similar to the periplasmic MBP concentration estimated for maltose-induced wild-type cells. Strains containing large deletions of the malB region and malT mutants lacking the positive regulator gene of the mal regulon also could be reconstituted for maltose chemotaxis, showing that no product of the mal regulon other than MBP is essential for maltose chemotaxis.

ATP-Binding Cassette Transporters↗

PubMed2006

Cellular and organ level localization of maltose in maltose-excess Arabidopsis mutants.

Maltose is the predominant form of carbon exported from the chloroplast at night. Plants that lack either the chloroplast maltose transporter or disproportionating enzyme 2 (DPE2, EC 2.4.1.25) have excess maltose in leaves. We confirmed that DPE2 is not associated with the chloroplast in Arabidopsis thaliana. Using non-aqueous fractionation methods, we found that essentially all the maltose in mex1-1 leaves is located inside chloroplasts but only 40% of maltose in dpe2-1 leaves is located inside chloroplasts. We found that maltose exists in a significant amount in the exudates collected from maltose-accumulating dpe2-1 Arabidopsis petioles. However, the amount of maltose in the exudates from mex1-1 petioles was not significantly different from that in wild-type phloem exudates. We found twice as much maltose in the roots of dpe2-1 plants relative to wild type but the maltose level in the roots of mex1-1 plants was not higher than wild type. We conclude that maltose accumulated in the cytosol of leaves can be carried from the shoots to the roots and that maltose accumulated in the chloroplast of mex1-1 leaves is not mobilized. By measuring the transcript levels and enzymatic activities, we show that maltose-metabolizing enzymes are active in wild-type roots. The amount of maltose moved from the shoots to the roots increased in dpe2-1 plants. The roots of dpe2-1 plants must have the capacity to metabolize the excess maltose.

Arabidopsis↗

PubMed2005

beta-Maltose is the metabolically active anomer of maltose during transitory starch degradation.

Maltose is the major form of carbon exported from the chloroplast at night as a result of transitory starch breakdown. Maltose exists as an alpha- or beta-anomer. We developed an enzymatic technique for distinguishing between the two anomers of maltose and tested the accuracy and specificity of this technique using beta-maltose liberated from maltoheptose by beta-amylase. This technique was used to investigate which form of maltose is present during transitory starch degradation in bean (Phaseolus vulgaris), wild-type Arabidopsis (Arabidopsis thaliana), two starch deficient Arabidopsis lines, and one starch-excess mutant of Arabidopsis. In Phaseolus and wild-type Arabidopsis, beta-maltose levels were low during the day but were much higher at night. In Arabidopsis plants unable to metabolize maltose due to a T-DNA insertion in the gene for the cytosolic amylomaltase, (Y. Lu, T.D. Sharkey [2004] Planta 218: 466-473) levels of alpha- and beta-maltose were high during both the day and night. In starchless mutants of Arabidopsis, total maltose levels were low and almost completely in the alpha-form. We also found that the subcellular concentration of beta-maltose at night was greater in the chloroplast than in the cytosol by 278 microm. We conclude that beta-maltose is the metabolically active anomer of maltose and that a sufficient gradient of beta-maltose exists between the chloroplast and cytosol to allow for passive transport of maltose out of chloroplasts at night.

Arabidopsis↗

PubMed1982

Maltose as an energy substrate during surgery: comparison of maltose with glucose.

To evaluate the usefulness of maltose as an energy substrate to be administered during surgery, five per cent maltose in lactated Ringer's solution and five per cent glucose in lactated Ringer's solution were administered to 10 cases each, at a rate of 5 ml . kg-1 hour (0.25 g . kg-1 hour as sugar) for two hours from the start of oral surgery, and their metabolic effects were compared. The maltose group showed a smaller increase in blood sugar level than the glucose group immediately after the completion of infusion. The mean plasma concentration of maltose reached a maximum of 121.6 mg/dl, and it remained at 12.3 mg/dl at four hours, indicating that the retention time of maltose in blood was longer than that of glucose. The mean recovery of sugar from four-hour urine samples was 3.26 per cent in the maltose group and 0.06 per cent in the glucose group respectively, showing greater urinary excretion by the maltose group. Plasma insulin was elevated less after maltose than after glucose infusion. The elevation following maltose infusion was considered not to be due to the administration of maltose per se, but to glucose produced from the maltose in the body. The anti-ketogenic effect of maltose was comparable to and tended to last longer than that of glucose. From overall assessment it was concluded that maltose exerts essentially the same metabolic effects as glucose when used under these conditions.

Adolescent↗

PubMed1982

Active transport of maltose in Escherichia coli K12. Role of the periplasmic maltose-binding protein and evidence for a substrate recognition site in the cytoplasmic membrane.

The active transport of maltose in Escherichia coli requires the products of five genes. These include a water-soluble periplasmic maltose-binding protein, three cytoplasmic membrane proteins, and an outer membrane protein. In order to evaluate the role of the maltose-binding protein in active transport, a nonpolar internal deletion of the structural gene for the maltose-binding protein was constructed. A strain which contains this deletion is unable to grow on maltose at an external concentration of 25 mM, even when the remaining components of the transport system are synthesized constitutively. This demonstrates that the periplasmic maltose-binding protein is essential for detectable translocation of maltose across the cytoplasmic membrane. Mal+ revertants of the deletion strain were obtained. In one of these strains, the remaining components of the maltose transport system gained the ability to translocate maltose across the membrane independently of the periplasmic binding protein. Maltose transport in this revertant strain is specific for maltose; it is not inhibited by other alpha and beta glucosides and galactosides. In contrast to the wild type, transport activity in the Mal+ revertant strain is retained by spheroplasts. The cytoplasmic membrane components of the maltose transport system in the revertant appear to form a substrate recognition site. It is likely that this site exists in wild type cells but is available only to substrate molecules that are bound to the maltose-binding protein. A model for the operation of the transport system is presented. In this model, the substrate recognition site in the cytoplasmic membrane is exposed to alternate sides of the membrane.

ATP-Binding Cassette Transporters↗

PubMed2004

Prolonged maltose-limited cultivation of Saccharomyces cerevisiae selects for cells with improved maltose affinity and hypersensitivity.

Prolonged cultivation (>25 generations) of Saccharomyces cerevisiae in aerobic, maltose-limited chemostat cultures led to profound physiological changes. Maltose hypersensitivity was observed when cells from prolonged cultivations were suddenly exposed to excess maltose. This substrate hypersensitivity was evident from massive cell lysis and loss of viability. During prolonged cultivation at a fixed specific growth rate, the affinity for the growth-limiting nutrient (i.e., maltose) increased, as evident from a decreasing residual maltose concentration. Furthermore, the capacity of maltose-dependent proton uptake increased up to 2.5-fold during prolonged cultivation. Genome-wide transcriptome analysis showed that the increased maltose transport capacity was not primarily due to increased transcript levels of maltose-permease genes upon prolonged cultivation. We propose that selection for improved substrate affinity (ratio of maximum substrate consumption rate and substrate saturation constant) in maltose-limited cultures leads to selection for cells with an increased capacity for maltose uptake. At the same time, the accumulative nature of maltose-proton symport in S. cerevisiae leads to unrestricted uptake when maltose-adapted cells are exposed to a substrate excess. These changes were retained after isolation of individual cell lines from the chemostat cultures and nonselective cultivation, indicating that mutations were involved. The observed trade-off between substrate affinity and substrate tolerance may be relevant for metabolic engineering and strain selection for utilization of substrates that are taken up by proton symport.

Biological Transport, Active↗

PubMed1992

Interaction between maltose-binding protein and the membrane-associated maltose transporter complex in Escherichia coli.

Active transport of maltose in Escherichia coli requires the presence of both maltose-binding protein (MBP) in the periplasm and a complex of MalF, MalG, and MalK proteins (FGK2) located in the cytoplasmic membrane. Earlier, mutants in malF or malG were isolated that are able to grow on maltose in the complete absence of MBP. When the wild-type malE+ allele, coding for MBP, was introduced into these MBP-independent mutants, they frequently lost their ability to grow on maltose. Furthermore, starting from these Mal- strains, Mal+ secondary mutants that contained suppressor mutations in malE were isolated. In this study, we examined the interaction of wild-type and mutant MBPs with wild-type and mutant FGK2 complexes by using right-side-out membrane vesicles. The vesicles from a MBP-independent mutant (malG511) transported maltose in the absence of MBP, with Km and Vmax values similar to those found in intact cells. However, addition of wild-type MBP to these mutant vesicles produced unexpected responses. Although malE+ malG511 cells could not utilize maltose, wild-type MBP at low concentrations stimulated the maltose uptake by malG511 vesicles. At higher concentrations of the wild-type MBP and maltose, however, maltose transport into malG511 vesicles became severely inhibited. This behaviour of the vesicles was also reflected in the phenotype of malE+ malG511 cells, which were found to be capable of transporting maltose from a low external concentration (1 microM), but apparently not from millimolar concentrations present in maltose minimal medium. We found that the mutant FGK2 complex, containing MalG511, had a much higher apparent affinity towards the wild-type MBP than did the wild-type FGK2 complex.(ABSTRACT TRUNCATED AT 250 WORDS)

ATP-Binding Cassette Transporters↗

PubMed1979

[Use of maltose and a mixture of maltose, fructose and xylitol in parenteral feeding].

Maltose or maltose in combination with fructose and xylitol was administered intravenously to eight healthy male subjects. Constant maltose levels could not be attained in the blood at an infusion rate of 0.125 g maltose/kg body-weight and hour. Maximal concentrations of maltose were found at the end of the infusion period. 8.6 +/- 1.2% of the administered radioactivity was excreted into urine within 8 hours. Regarding the enzymatically determined maltose and glucose, the maltose balance was more favorable with the loss of only 3--3.5% carbohydrates into urine. The highest oxidation rate of the administered maltose was 1.5 g maltose/human volunteer and hour. During the experimental period of 8 hours 7.4 g maltose, corresponding to 37% of the applied dosis of the disaccharid, has been oxidized to and excreted as 14CO2. Xylitol and fructose did not effect utilisation and balance of maltose. Only the urinary excretion of glucose was higher when the combined solution was applied. At a limited infusion rate (0.125 g maltose/kg body-weight and hour) maltose or the combined solution maltose--xylitol--fructose may be recommended for parenteral nutrition.

Adult↗

PubMed1993

Maltose and maltotriose can be formed endogenously in Escherichia coli from glucose and glucose-1-phosphate independently of enzymes of the maltose system.

The maltose system in Escherichia coli consists of cell envelope-associated proteins and enzymes that catalyze the uptake and utilization of maltose and alpha,1-4-linked maltodextrins. The presence of these sugars in the growth medium induces the maltose system (exogenous induction), even though only maltotriose has been identified in vitro as an inducer (O. Raibaud and E. Richet, J. Bacteriol., 169:3059-3061, 1987). Induction is dependent on MalT, the positive regulator protein of the system. In the presence of exogenous glucose, the maltose system is normally repressed because of catabolite repression and inducer exclusion brought about by the phosphotransferase-mediated vectorial phosphorylation of glucose. In contrast, the increase of free, unphosphorylated glucose in the cell induces the maltose system. A ptsG ptsM glk mutant which cannot grow on glucose can accumulate [14C]glucose via galactose permeases. In this strain, internal glucose is polymerized to maltose, maltotriose, and maltodextrins in which only the reducing glucose residue is labeled. This polymerization does not require maltose enzymes, since it still occurs in malT mutants. Formation of maltodextrins from external glucose as well as induction of the maltose system is absent in a mutant lacking phosphoglucomutase, and induction by external glucose could be regained by the addition of glucose-1-phosphate entering the cells via a constitutive glucose phosphate transport system. malQ mutants, which lack amylomaltase, are constitutive for the expression of the maltose genes. This constitutive nature is due to the formation of maltose and maltodextrins from the degradation of glycogen.

Adenosine Diphosphate↗

PubMed1991

The malX malY operon of Escherichia coli encodes a novel enzyme II of the phosphotransferase system recognizing glucose and maltose and an enzyme abolishing the endogenous induction of the maltose system.

Mutants lacking MalK, a subunit of the binding protein-dependent maltose-maltodextrin transport system, constitutively express the maltose genes. A second site mutation in malI abolishes the constitutive expression. The malI gene (at 36 min on the linkage map) codes for a typical repressor protein that is homologous to the Escherichia coli LacI, GalR, or CytR repressor (J. Reidl, K. Römisch, M. Ehrmann, and W. Boos, J. Bacteriol. 171:4888-4899, 1989). We now report that MalI regulates an adjacent and divergently oriented operon containing malX and malY. MalX encodes a protein with a molecular weight of 56,654, and the deduced amino acid sequence of MalX exhibits 34.9% identity to the enzyme II of the phosphototransferase system for glucose (ptsG) and 32.1% identity to the enzyme II for N-acetylglucosamine (nagE). When constitutively expressed, malX can complement a ptsG ptsM double mutant for growth on glucose. Also, a delta malE malT(Con) strain that is unable to grow on maltose due to its maltose transport defect becomes Mal+ after introduction of malI::Tn10 and the plasmid carrying malX. MalX-mediated transport of glucose and maltose is likely to occur by facilitated diffusion. We conclude that malX encodes a phosphotransferase system enzyme II that can recognize glucose and maltose as substrates even though these sugars may not represent the natural substrates of the system. The second gene in the operon, malY, encodes a protein of 43,500 daltons. Its deduced amino acid sequence exhibits weak homology to aminotransferase sequences. The presence of plasmid-encoded MalX alone was sufficient for complementing growth on glucose in a ptsM ptsG glk mutant, and the plasmid-encoded MalY alone was sufficient to abolish the constitutivity of the mal genes in a malK mutant. The overexpression of malY in a strain that is wild type with respect to the maltose genes strongly interferes with growth on maltose. This is not the case in a malT(Con) strain that expresses the mal genes constitutively. We conclude that malY encodes an enzyme that degrades the inducer of the maltose system or prevents its synthesis.

Amino Acid Sequence↗

PubMed2000

Crystal transformation from anhydrous alpha-maltose to hydrous beta-maltose and from anhydrous trehalose to hydrous trehalose.

Anhydrous sugars such as maltose and trehalose are useful for making dry powder of foods and liquids. The crystal-transformation rate of maltose and trehalose were investigated under humid conditions and by kneading. The enthalpy for solubilization was 7.0 kJ/mol for the anhydrous maltose. The crystal-transformation rate of anhydrous alpha-maltose to hydrous beta-maltose depended on the temperature at 75% humidity. However, that of anhydrous trehalose did not depend on the temperature, and transformation was very rapid. An anomeric change to maltose and no such change to trehalose might have caused this. The activation energy of crystal transformation was 79 kJ/mol for maltose and zero for trehalose. The rate of crystal transformation of anhydrous maltose while kneading depended on the purity of the anhydrous alpha-maltose and the amount of water present. This crystal transformation rate fitted the Avrami equation.

Crystallization↗

PubMed1994

Maltose transport in Aeromonas hydrophila: purification, biochemical characterization and partial protein sequence analysis of a periplasmic maltose-binding protein.

A clinical isolate of Aeromonas hydrophila was demonstrated to transport [14C]maltose with similar kinetics to enteric bacteria (Km: 0.3 microM; Vmax: 22 nmol min-1 per 10(9) cells). The uptake of [14C]maltose was completely inhibited in the presence of unlabelled maltose or maltodextrins, whereas other mono- and disaccharides, such as glucose, galactose, sucrose, lactose or melibiose, had no effect. A protein with an apparent molecular mass of 39 kDa (maltose-binding protein; MBP) was identified in osmotic-shock fluid of maltose-grown cells by SDS-gel electrophoresis, and was purified to homogeneity by either amylose affinity chromatography or ion-exchange chromatography. Equilibrium dialysis experiments revealed the ability of the purified protein to bind [14C]maltose with high affinity (KD = 1.6 microM). Unlabelled maltose and maltodextrins competed for the binding site. In a reconstitution experiment, A. hydrophila MBP poorly restored the transport activity of a binding-protein-deficient Escherichia coli (delta malE) mutant. N-terminal sequence analyses of the purified native protein and of peptides generated by cleavage with CNBr and subsequently separated by HPLC revealed about 56% identical amino acid residues, as compared to enterobacterial MBPs. We conclude that maltose is transported into A. hydrophila via a binding-protein-dependent transport system.

ATP-Binding Cassette Transporters↗

PubMed1985

Interspecific reconstitution of maltose transport and chemotaxis in Escherichia coli with maltose-binding protein from various enteric bacteria.

In Escherichia coli, the periplasmic maltose-binding protein (MBP), the product of the malE gene, is the primary recognition component of the transport system for maltose and maltodextrins. It is also the maltose chemoreceptor, in which capacity it interacts with the signal transducer Tar (taxis to aspartate and some repellents). In studies of the maltose system in other members of the family Enterobacteriaceae, we found that MBP is produced by Salmonella typhimurium, Klebsiella pneumoniae, Enterobacter aerogenes, and Serratia marcescens. MBP from all of these species cross-reacted with antibody against the E. coli protein and had a similar molecular weight (about 40,000). The Shigella flexneri and Proteus mirabilis strains we examined did not synthesize MBP. The isoelectric points of MBP from different species varied from the acid extreme of E. coli (4.8) to the basic extreme of E. aerogenes (8.9). All species with MBP transported maltose with high affinity, although the Vmax for K. pneumoniae was severalfold lower than that for the other species. Maltose chemotaxis was observed only in E. coli and E. aerogenes. In S. typhimurium LT2, Tar was completely inactive in maltose taxis, although it signaled normally in response to aspartate. MBP isolated from all five species could be used to reconstitute maltose transport and taxis in a delta malE strain of E. coli after permeabilization of the outer membrane with calcium.

ATP-Binding Cassette Transporters↗

PubMed2000

Maltose and maltodextrin transport in the thermoacidophilic gram-positive bacterium Alicyclobacillus acidocaldarius is mediated by a high-affinity transport system that includes a maltose binding protein tolerant to low pH.

We have studied the uptake of maltose in the thermoacidophilic gram-positive bacterium Alicyclobacillus acidocaldarius, which grows best at 57 degrees C and pH 3.5. Under these conditions, accumulation of [(14)C]maltose was observed in cells grown with maltose but not in those grown with glucose. At lower temperatures or higher pH values, the transport rates substantially decreased. Uptake of radiolabeled maltose was inhibited by maltotetraose, acarbose, and cyclodextrins but not by lactose, sucrose, or trehalose. The kinetic parameters (K(m) of 0.91 +/- 0.06 microM and V(max) ranging from 0.6 to 3.7 nmol/min/mg of protein) are consistent with a binding protein-dependent ATP binding cassette (ABC) transporter. A corresponding binding protein (MalE) that interacts with maltose with high affinity (K(d) of 1.5 microM) was purified from the culture supernatant of maltose-grown cells. Immunoelectron microscopy revealed distribution of the protein throughout the cell wall. The malE gene was cloned and sequenced. Five additional open reading frames, encoding components of a maltose transport system (MalF and MalG), a putative transcriptional regulator (MalR), a cyclodextrinase (CdaA), and an alpha-glucosidase (GlcA), were identified downstream of malE. The malE gene lacking the DNA sequence that encodes the signal sequence was expressed in Escherichia coli. The purified wild-type and recombinant proteins bind maltose with high affinity over a wide pH range (2.5 to 7) and up to 80 degrees C. Recombinant MalE cross-reacted with an antiserum raised against the wild-type protein, thereby indicating that the latter is the product of the malE gene. The MalE protein might be well suited as a model to study tolerance of proteins to low pH.

ATP-Binding Cassette Transporters↗

PubMed1990

Substrate-induced activation of maltose phosphorylase: interaction with the anomeric hydroxyl group of alpha-maltose and alpha-D-glucose controls the enzyme's glucosyltransferase activity.

Maltose phosphorylase, long considered strictly specific for beta-D-glucopyranosyl phosphate (beta-D-glucose 1-P), was found to catalyze the reaction beta-D-glucosyl fluoride + alpha-D-glucose----alpha-maltose + HF, at a rapid rate, V = 11.2 +/- 1.2 mumol/(min.mg), and K = 13.1 +/- 4.4 mM with alpha-D-glucose saturating, at 0 degrees C. This reaction is analogous to the synthesis of maltose from beta-D-glucose 1-P + D-glucose (the reverse of maltose phosphorolysis). In acting upon beta-D-glucosyl fluoride, maltose phosphorylase was found to use alpha-D-glucose as a cosubstrate but not beta-D-glucose or other close analogs (e.g., alpha-D-glucosyl fluoride) lacking an axial 1-OH group. Similarly, the enzyme was shown to use alpha-maltose as a substrate but not beta-maltose or close analogs (e.g., alpha-maltosyl fluoride) lacking an axial 1-OH group. These results indicate that interaction of the axial 1-OH group of the disaccharide donor or sugar acceptor with a particular protein group near the reaction center is required for effective catalysis. This interaction appears to be the means that leads maltose phosphorylase to promote a narrowly defined set of glucosyl transfer reactions with little hydrolysis, in contrast to other glycosylases that catalyze both hydrolytic and nonhydrolytic reactions.

Binding Sites↗

PubMed2005

Clustering of MAL genes in Hansenula polymorpha: cloning of the maltose permease gene and expression from the divergent intergenic region between the maltose permease and maltase genes.

Hansenula polymorpha uses maltase to grow on maltose and sucrose. Inspection of genomic clones of H. polymorpha showed that the maltase gene HPMAL1 is clustered with genes corresponding to Saccharomyces cerevisiae maltose permeases and MAL activator genes orthologues. We sequenced the H. polymorpha maltose permease gene HPMAL2 of the cluster. The protein (582 amino acids) deduced from the HPMAL2 gene is predicted to have eleven transmembrane domains and shows 39-57% identity with yeast maltose permeases. The identity of the protein is highest with maltose permeases of Debaryomyces hansenii and Candida albicans. Expression of the HPMAL2 in a S. cerevisiae maltose permease-negative mutant CMY1050 proved functionality of the permease protein encoded by the gene. HPMAL1 and HPMAL2 genes are divergently positioned similarly to maltase and maltose permease genes in many yeasts. A two-reporter assay of the expression from the HPMAL1-HPMAL2 intergenic region showed that expression of both genes is coordinately regulated, repressed by glucose, induced by maltose, and that basal expression is higher in the direction of the permease gene.

Amino Acid Sequence↗

PubMed1987

Silent and functional changes in the periplasmic maltose-binding protein of Escherichia coli K12. I. Transport of maltose.

The malE gene encodes the periplasmic maltose-binding protein (MBP). Nineteen mutations that still permit synthesis of stable MBP were generated by random insertion of a BamHI octanucleotide into malE and six additional mutations by in-vitro recombinations between mutant genes. The sequence changes were determined; in most cases the linker insertion is accompanied by a small deletion (30 base-pairs on average). The mutant MBP were studied for export, growth on maltose and maltodextrins, maltose transport and binding, and maltose-induced fluorescence changes. Sixteen mutant MBP (out of 21 studied in detail) were found in the periplasmic space: 12 of them retained a high affinity for maltose, and 10 activity for growth on maltose. The results show that several regions of MBP are dispensable for stability, substrate binding and export. Three regions (residues 207 to 220, 297 to 303 and 364 to 370) may be involved in interactions with the MalF or MalG proteins. A region near the C-terminal end is important for maltose binding. Two regions of the mature protein (residues 18 to 42 and 280 to 296) are required for export to, or solubility in, the periplasm.

ATP-Binding Cassette Transporters↗

PubMed1985

Dependence of maltose transport and chemotaxis on the amount of maltose-binding protein.

Maltose-binding protein (MBP) is essential for maltose transport and chemotaxis in Escherichia coli. To perform these functions it must interact with two sets of cytoplasmic membrane proteins, the MalFGK transport complex and the chemotactic signal transducer Tar. MBP is present at high concentrations, on the order of 1 mM, in the periplasm of maltose-induced or malTc constitutive cells. To determine how the amount of MBP affects transport and taxis, we utilized a series of malE signal-sequence mutations that interfere with export of MBP. The MBP content in shock fluid from cells carrying the various mutations ranged from 4 to 23% of the malE+ level. The apparent Km for maltose transport varied by less than a factor of 2 among malE+ and mutant strains. At a saturating maltose concentration 9% (approximately 90 microM) of the malE+ amount of MBP was required for half-maximal uptake rates. Transport exhibited a sigmoidal dependence on the amount of periplasmic MBP, indicating that MBP may be involved in a cooperative interaction at some stage of the transport process. The chemotactic response to a saturating maltose stimulus exhibited a first-order dependence on the amount of periplasmic MBP. Thus, interaction of a single substrate-bound MBP with Tar appears sufficient to initiate a chemotactic signal from the transducer. A half-maximal chemotactic response occurred at 25% of the malE+ MBP level, suggesting that in vivo the KD for binding of maltose-loaded MBP to Tar is quite high (approximately 250 microM).

ATP-Binding Cassette Transporters↗