arbutin and salicin

Bacteria have evolved various mechanisms to extract utilizable substrates from available resources and consequently acquire fitness advantage over competitors. One of the strategies is the exploitation of cryptic cellular functions encoded by genetic systems that are silent under laboratory conditions, such as the bgl (β-glucoside) operon of E. coli. The bgl operon of Escherichia coli, involved in the uptake and utilization of aromatic β-glucosides salicin and arbutin, is maintained in a silent state in the wild type organism by the presence of structural elements in the regulatory region. This operon can be activated by mutations that disrupt these negative elements. The fact that the silent bgl operon is retained without accumulating deleterious mutations seems paradoxical from an evolutionary view point. Although this operon appears to be silent, specific physiological conditions might be able to regulate its expression and/or the operon might be carrying out function(s) apart from the utilization of aromatic β-glucosides. This is consistent with the observations that the activated operon confers a Growth Advantage in Stationary Phase (GASP) phenotype to Bgl(+) cells and exerts its regulation on at least twelve downstream target genes.

Topics: Arbutin; Benzyl Alcohols; beta-Glucosidase; Escherichia coli; Gene Expression Regulation; Glucosides; Operon

The cryptic β-glucoside GFB (

Topics: Arbutin; Bacterial Proteins; Bacteriological Techniques; Benzyl Alcohols; Culture Media; DNA Transposable Elements; Escherichia coli; Gene Expression Regulation, Bacterial; Glucosides; Mutagenesis, Insertional; Operon; RNA-Binding Motifs; RNA-Binding Proteins

Salicylic acid (SA) is a plant hormone involved in a number of physiological responses including both local and systemic resistance of plants to pathogens. In Arabidopsis, SA is glucosylated to form either SA 2-O-β-d-glucose (SAG) or SA glucose ester (SGE). In this study, we show that SAG accumulates in the vacuole of Arabidopsis, while the majority of SGE was located outside the vacuole. The uptake of SAG by vacuolar membrane-enriched vesicles isolated from Arabidopsis was stimulated by the addition of MgATP and was inhibited by both vanadate (ABC transporter inhibitor) and bafilomycin A

Topics: Adenosine Triphosphate; Arabidopsis; Arbutin; ATP-Binding Cassette Transporters; Benzyl Alcohols; Carbon Radioisotopes; Chromatography, High Pressure Liquid; Glucose; Glucosides; Gramicidin; Intracellular Membranes; Kinetics; Metabolome; Protoplasts; Quercetin; Salicylic Acid; Time Factors; Transport Vesicles; Vacuolar Proton-Translocating ATPases; Vacuoles; Verapamil

Topics: Arbutin; Benzyl Alcohols; Cellobiose; DNA Transposable Elements; Escherichia coli; Escherichia coli Proteins; Evolution, Molecular; Glucosides; Mutation; Operon; Repressor Proteins; Shigella sonnei

The total synthesis of two natural phenolglycosides of the family Salicaceae, namely: populoside and 2-(β-d-glucopyranosyloxy)-5-hydroxy benzyl (3-methoxy-4-hydroxy) cinnamoate and nine not found yet in plants acyl derivatives of phenoglycosides: 2-(β-d-glucopyranosyloxy)-benzylcinnamoate, 2-(β-d-glucopyranosyloxy)-benzyl (4-hydroxy) benzoate, 2-(β-d-glucopyranosyloxy)-benzyl (3-methoxy-4-hydroxy) benzoate, 2-(β-d-glucopyranosyloxy)-5-hydroxy benzyl (3,4-dihydroxy) cinnamoate, 2-(β-d-glucopyranosyloxy)-5-hydroxy benzylcinnamoate, 2-(β-d-glucopyranosyloxy)-5-hydroxy benzyl (4-hydroxy) benzoate, 2-(β-d-glucopyranosyloxy)-5-hydroxy benzyl (3-methoxy-4-hydroxy) benzoate, 2-(β-d-glucopyranosyloxy)-5-benzoyloxy benzylbenzoate and 4-(β-d-glucopyranosyloxy)-phenylbenzoate, starting from readily available phenols and glucose was developed for the first time.

Topics: Arbutin; Benzyl Alcohols; Benzyl Compounds; Cinnamates; Glucose; Glucosides; Hydroquinones; Phenols

Utilization of the aryl-β-glucosides salicin or arbutin in most wild-type strains of E. coli is achieved by a single-step mutational activation of the bgl operon. Shigella sonnei, a branch of the diverse E. coli strain tree, requires two sequential mutational steps for achieving salicin utilization as the bglB gene, encoding the phospho-β-glucosidase B, harbors an inactivating insertion. We show that in a natural isolate of S. sonnei, transcriptional activation of the gene SSO1595, encoding a phospho-β-glucosidase, enables salicin utilization with the permease function being provided by the activated bgl operon. SSO1595 is absent in most commensal strains of E. coli, but is present in extra-intestinal pathogens as bgcA, a component of the bgc operon that enables β-glucoside utilization at low temperature. Salicin utilization in an E. coli bglB laboratory strain also requires a two-step activation process leading to expression of BglF, the PTS-associated permease encoded by the bgl operon and AscB, the phospho-β-glucosidase B encoded by the silent asc operon. BglF function is needed since AscF is unable to transport β-glucosides as it lacks the IIA domain involved in phopho-relay. Activation of the asc operon in the Sal(+) mutant is by a promoter-up mutation and the activated operon is subject to induction. The pathway to achieve salicin utilization is therefore diverse in these two evolutionarily related organisms; however, both show cooperation between two silent genetic systems to achieve a new metabolic capability under selection.

Topics: Arbutin; Benzyl Alcohols; DNA, Bacterial; Escherichia coli; Evolution, Molecular; Gene Expression Regulation, Bacterial; Gene Silencing; Genes, Bacterial; Glucosides; Mutation; Operon; Shigella sonnei; Transcription Initiation Site; Transcriptional Activation

Escherichia coli strains, in general, do not ferment cellobiose and aryl-beta-D-glucosidic sugars, although "cryptic" beta-d-glucoside systems have been characterized. Here we describe an additional cryptic operon (bgc) for the utilization of cellobiose and the aryl-beta-d-glucosides arbutin and salicin at low temperature. The bgc operon was identified by the characterization of beta-glucoside-positive mutants of an E. coli septicemia strain (i484) in which the well-studied bgl (aryl-beta-d-glucoside) operon was deleted. These bgc* mutants appeared after 5 days of incubation on salicin indicator plates at 28 degrees C. The bgc operon codes for proteins homologous to beta-glucoside/cellobiose-specific phosphoenolpyruvate-dependent phosphotransfer system permease subunits IIB (BgcE), IIC (BgcF), and IIA (BgcI); a porin (BgcH); and a phospho-beta-D-glucosidase (BgcA). Next to the bgc operon maps the divergent bgcR gene, which encodes a GntR-type transcriptional regulator. Expression of the bgc operon is dependent on the cyclic-AMP-dependent regulator protein CRP and positively controlled by BgcR. In the bgc* mutants, a single nucleotide exchange enhances the activity of the bgc promoter, rendering it BgcR independent. Typing of a representative collection of E. coli demonstrated the prevalence of bgc in strains of phylogenetic group B2, representing mainly extraintestinal pathogens, while it is rare among commensal E. coli strains. The bgc locus is also present in the closely related species Escherichia albertii. Further, bioinformatic analyses demonstrated that homologs of the bgc genes exist in the enterobacterial Klebsiella, Enterobacter, and Citrobacter spp. and also in gram-positive bacteria, indicative of horizontal gene transfer events.

Topics: Arbutin; Benzyl Alcohols; beta-Glucosidase; Cellobiose; Citrobacter; Cyclic AMP Receptor Protein; Enterobacter; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Glucosides; Klebsiella; Operon; Phylogeny; Repressor Proteins; Sepsis; Sequence Homology; Synteny; Urinary Tract Infections

The phosphoenolpyruvate : carbohydrate phosphotransferase system (PTS) catalyses carbohydrate transport by coupling it to phosphorylation. Previously, we reported a Corynebacterium glutamicum R beta-glucoside PTS encoded by bglF. Here we report that C. glutamicum R contains an additional beta-glucoside PTS gene, bglF2, organized in a cluster with a putative phospho-beta-glucosidase gene, bglA2, and a putative antiterminator, bglG2. While single gene disruption strains of either bglF or bglF2 were able to utilize salicin or arbutin as sole carbon sources, a double disruption strain exhibited defects in utilization of both carbon sources. Expression of both bglF and bglF2 was induced in the presence of either salicin or arbutin, although disruption of bglG2 affected only bglF2 expression. Moreover, in the presence of either salicin or arbutin, glucose completely repressed the expression of bglF but only slightly repressed that of bglF2. We conclude that BglF and BglF2 have a redundant role in beta-glucoside transport even though the catabolite repression control of their encoding genes is different. We also show that expression of both bglF and bglF2 requires the general PTS.

Topics: Arbutin; Bacterial Proteins; Base Sequence; Benzyl Alcohols; Corynebacterium glutamicum; Gene Expression Regulation, Bacterial; Genes, Bacterial; Glucose; Glucosides; Molecular Sequence Data; Multigene Family; Phosphoenolpyruvate Sugar Phosphotransferase System; RNA, Bacterial

AtSUC9 (At5g06170), a sucrose (Suc) transporter from Arabidopsis (Arabidopsis thaliana) L. Heynh., was expressed in Xenopus (Xenopus laevis) oocytes, and transport activity was analyzed. Compared to all other Suc transporters, AtSUC9 had an ultrahigh affinity for Suc (K(0.5) = 0.066 +/- 0.025 mm). AtSUC9 showed low substrate specificity, similar to AtSUC2 (At1g22710), and transported a wide range of glucosides, including helicin, salicin, arbutin, maltose, fraxin, esculin, turanose, and alpha-methyl-d-glucose. The ability of AtSUC9 to transport 10 glucosides was compared directly with that of AtSUC2, HvSUT1 (from barley [Hordeum vulgare]), and ShSUT1 (from sugarcane [Saccharum hybrid]), and results indicate that type I and type II Suc transporters have different substrate specificities. AtSUC9 protein was localized to the plasma membrane by transient expression in onion (Allium cepa) epidermis. Using a whole-gene translational fusion to beta-glucuronidase, AtSUC9 expression was found in sink tissues throughout the shoots and in flowers. AtSUC9 expression in Arabidopsis was dependent on intragenic sequence, and this was found to also be true for AtSUC1 (At1g71880) but not AtSUC2. Plants containing mutations in Suc transporter gene AtSUC9 were found to have an early flowering phenotype under short-day conditions. The transport properties of AtSUC9 indicate that it is uniquely suited to provide cellular uptake of Suc at very low extracellular Suc concentrations. The mutant phenotype of atsuc9 alleles indicates that AtSUC9 activity leads to a delay in floral transition.

Topics: Animals; Arabidopsis; Arabidopsis Proteins; Arbutin; Benzyl Alcohols; Biological Transport; Cell Membrane; Flowers; Gene Expression Regulation, Plant; Glucosides; Hydrogen-Ion Concentration; Membrane Transport Proteins; Mutation; Phenotype; Plant Proteins; Regulatory Elements, Transcriptional; Substrate Specificity; Sucrose; Xenopus

The genome of Streptococcus mutans UA159 contains two phospho-beta-glucosidase genes, bglA and celA, which occur in operon-like arrangements along with genes for components of phosphotransferase transport systems and a third phospho-beta-glucosidase encoded by the arb gene, which does not have its own associated transport system but relies on uptake by the bgl or cel systems. Targeted inactivation of each of the phospho-beta-glucosidase genes revealed that bglA is involved in aesculin hydrolysis, celA is essential for utilisation of cellobiose, amygdalin, gentobiose and salicin, and arb is required for utilisation of arbutin. Inactivation of genes for the phosphotransferase systems revealed an overlap of specificity for transport of beta-glucosides and also indicated that further, unidentified transport systems exist. The cel and arb genes are subject to catabolite repression by glucose, but the regM gene is not essential for catabolite repression. Screening a collection of isolates of S. mutans revealed strains with deletions affecting the msm, bgl and/or cel operons. The phenotypes of these strains could largely be explained on the basis of the results obtained from the knockout mutants of S. mutans UA159 but also indicated the existence of other pathways apparently absent from UA159. The extensive genetic and phenotypic variation found in beta-glucoside metabolism indicates that there may be extensive heterogeneity in the species.

Topics: Amygdalin; Arbutin; Bacterial Proteins; Benzyl Alcohols; beta-Glucosidase; Cellobiose; Cellulase; Esculin; Gene Deletion; Gene Silencing; Genetic Variation; Genome, Bacterial; Glucosides; Humans; Hydrolysis; Mutation; Operon; Phenotype; Phosphoenolpyruvate Sugar Phosphotransferase System; Streptococcus mutans

The paper aimed to transform exogenous precursors with in vitro cultures of Datura meteloides, Coronilla varia, Leuzea carthamoides and Schisandra chinensis. These cultures were added the precursors of arbutin and salicin (phenylalanine, cinnamic, p-coumaric, p-anisoic, o-coumaric, salicylic acids, salicylaldehyde, helicin), not yet tested by the present authors. The culture of Schisandra chinensis was also added, besides the above-mentioned precursors, hydroquinone, because this culture had not been employed for biotransformation purposes yet. The precursors tested were used in a concentration of 100 mg x l(-1) and the period of their action was 6; 12; 24; 48, and 168 hours. Positive results (both TLC and HPLC) in arbutin production were obtained in the culture of Schisandra chinensis after an addition of hydroquinone. The largest amount of arbutin in callus cultures was measured after a week's cultivation with hydroquinone (5.08 %). In this experimental variant, arbutin was released also to the culture medium. Our results revealed salicylaldehyde to be the optimal precursor of salicin. It was transformed by the culture of Datura meteloides after 6; 24, and 168 hours and by the culture of Coronilla varia after 6 hours. In comparison with arbutin, its amount was smaller.

Topics: Arbutin; Benzyl Alcohols; Biotransformation; Cell Culture Techniques; Glucosides; Plants, Medicinal

During periods of human expansion into new environments, recognition of bitter natural toxins through taste may have conferred an important selective advantage. The G protein-coupled receptor encoded by TAS2R16 mediates response to salicin, amygdalin, and many bitter beta-glucopyranosides. beta-glucopyranosides are ubiquitous in nature, with many having a highly toxic cyanogenic activity.. We examined evidence for natural selection on the human receptor TAS2R16 by sequencing the entire coding region, as well as part of the 5' and 3' UTRs, in 997 individuals from 60 human populations. We detected signatures of positive selection, indicated by an excess of evolutionarily derived alleles at the nonsynonymous site K172N and two linked sites and significant values of Fay and Wu's H statistics in 19 populations. The estimated age range for the common ancestor of the derived N172 variant is 78,700-791,000 years, placing it in the Middle Pleistocene and before the expansion of early humans out of Africa. Using calcium imaging in cells expressing different receptor variants, we showed that N172 is associated with an increased sensitivity to salicin, arbutin, and five different cyanogenic glycosides.. We have detected a clear signal of positive selection at the bitter-taste receptor gene TAS2R16. We speculate that the increased sensitivity that is shown toward harmful cyanogenic glycosides and conferred by the N172 allele may have driven the signal of selection at an early stage of human evolution.

Topics: Alleles; Amygdalin; Arbutin; Base Sequence; Benzyl Alcohols; Evolution, Molecular; Genetic Variation; Genetics, Population; Glucosides; Glycosides; Haplotypes; Humans; Immunohistochemistry; Molecular Sequence Data; Receptors, G-Protein-Coupled; Selection, Genetic; Sequence Analysis, DNA

Four aryl-phospho-beta- d-glucosidases were identified in Bacillus subtilis by using 4-methylumbelliferyl-phospho-beta- d-glucopyranoside as a substrate. Two of these enzymes are the products of the bglA and bglH genes, previously suggested to encode aryl-phospho-beta- d-glucosidases, while the other enzymes are encoded by the yckE and ydhP genes. Together, these four genes account for >99.9% of the glucosidase activity in B. subtilis on aryl-phospho-beta- d-glucosides. yckE was expressed at a low and constant level during growth, sporulation, and spore germination, and was not induced by aryl-beta- d-glucosides. ydhP was also not induced by aryl-beta- d-glucosides. However, while ydhP was expressed at only a very low level in exponential-phase cells and germinating spores, this gene was expressed at a higher levels upon entry into the stationary phase of growth. Strains lacking yckE or ydhP exhibited no defects in growth, sporulation, or spore germination or in growth on aryl-beta- d-glucosides. However, a strain lacking bglA, bglH and yckE grew poorly if at all on aryl-beta- d-glucosides as the sole carbon source.

Topics: Arbutin; Bacillus subtilis; Benzyl Alcohols; Gene Expression Regulation, Bacterial; Genes, Bacterial; Glucose; Glucosidases; Glucosides; Hymecromone; Spores, Bacterial

Previous studies have suggested that carbohydrates may affect expression of virulence genes in Listeria monocytogenes. Which carbohydrates influence virulence gene expression and how carbohydrates mediate expression, however, is not clear. The goal of this work was to examine how carbohydrates affect virulence gene expression in L. monocytogenes 10403S. Growth studies were conducted in medium containing glucose and various sugars. Metabolism of arbutin, arabitol, cellobiose, mannose, maltose, trehalose, and salicin were repressed in the presence of glucose. Only when glucose was consumed were these sugars fermented, indicating that catabolite repression by glucose had occurred. To determine whether virulence gene expression was also influenced by catabolite repression, we performed primer extension experiments, using primers for hly and prfA, which encode for a hemolysin and the regulator protein PrfA, respectively. In the presence of cellobiose and arbutin, transcription of hemolysin was reduced. However, none of the sugars affected transcription of prfA. The results demonstrate that catabolite repression occurs in L. monocytogenes and suggests that, at least in strain 10403S, cellobiose and arbutin repress expression of hemolysin.

Topics: Arbutin; Bacterial Proteins; Bacterial Toxins; Benzyl Alcohols; Carbohydrate Metabolism; Cellobiose; Fermentation; Gene Expression Regulation, Bacterial; Glucose; Glucosides; Heat-Shock Proteins; Hemolysin Proteins; Listeria monocytogenes; Maltose; Mannose; Peptide Termination Factors; Sugar Alcohols; Trans-Activators; Transcription, Genetic; Trehalose; Virulence Factors

The Arabidopsis sucrose transporter AtSUC2 is expressed in the companion cells of the phloem (specialized vascular tissue) and is essential for the long distance transport of carbohydrates within the plant. A variety of glucosides are known to inhibit sucrose uptake into yeast expressing AtSUC2; however, it remains unknown whether glucosides other than sucrose could serve as transported substrates. By expression of AtSUC2 in Xenopus oocytes and two-electrode voltage clamping, we have tested the ability of AtSUC2 to transport a range of physiological and synthetic glucosides. Sucrose induced inward currents with a K0.5 of 1.44 mM at pH 5 and a membrane potential of -137 mV. Of the 24 additional sugars tested, 8 glucosides induced large inward currents allowing kinetic analysis. These glucosides were maltose, arbutin (hydroquinone-beta-D-glucoside), salicin (2-(hydroxymethyl)phenyl-beta-D-glucoside), alpha-phenylglucoside, beta-phenylglucoside, alpha-paranitrophenylglucoside, beta-paranitrophenylglucoside, and paranitrophenyl-beta-thioglucoside. In addition, turanose and alpha-methylglucoside induced small but significant inward currents indicating that they were transported by At-SUC2. The results indicate that AtSUC2 is not highly selective for alpha-over beta-glucosides and may function in transporting glucosides besides sucrose into the phloem, and the results provide insight into the structural requirements for transport by AtSUC2.

Topics: Animals; Arabidopsis Proteins; Arbutin; Benzyl Alcohols; Carbohydrate Conformation; Electric Conductivity; Female; Gene Expression; Glucosides; Kinetics; Maltose; Membrane Potentials; Membrane Transport Proteins; Oocytes; Plant Proteins; Sucrose; Transfection; Xenopus laevis

The present study characterised 73 Hafnia alvei isolates and five Escherichia isolates (originally identified as H. alvei) isolated from cases of diarrhoeal disease by the International Centre for Diarrhoeal Disease Research Branch (ICDDRB) in Bangladesh. Based upon the hydrolysis of arbutin and aesculin and the fermentation of salicin and D-arabinose, four distinct biotypes could be recognised among the 73 H. alvei isolates tested; biotype 1 (D-(-)-arabinose-positive only) accounted for 75% of all isolates analysed. Hydrolysis of aglycone compounds such as arbutin, salicin and aesculin appeared to be associated with expression of beta-glucosidase activity. ICDDRB isolates, when compared with type or reference strains of H. alvei, were shown not to belong to the genus Hafnia based upon resistance to Hafnia-specific bacteriophage 1672, possession of the phoE gene, expression of glutamate decarboxylase activity and significant 16S rDNA sequence divergence (approximately 8%) from the type strain, ATCC 13337T. True H. alvei strains, implicated in outbreaks of diarrhoeal disease in Canada, lacked the eaeA gene in contrast to ICDDRB isolates. Twenty-two H. alvei isolates were selected for further study. Based upon partial 16S rDNA sequencing, these 22 isolates fell into two genomic groups (genomospecies), identical to DNA groups previously established by DNA hybridisation studies. Markers such as motility, biotype, or enzymic or carbohydrate fermentation patterns did not correlate totally with DNA grouping, although malonate utilisation appeared to be the single best discriminatory phenotype. The results indicate that the genus Hafnia is heterogeneous and there do not appear to be any laboratory data available specifically linking these organisms to gastro-enteritis.

Topics: Adhesins, Bacterial; Animals; Arabinose; Arbutin; Bacterial Typing Techniques; Bangladesh; Benzyl Alcohols; beta-Glucosidase; Carrier Proteins; Diarrhea; Electrophoresis, Gel, Pulsed-Field; Escherichia coli; Escherichia coli Proteins; Esculin; Fermentation; Genes, Bacterial; Genotype; Glucosides; Hafnia alvei; Humans; Hydrolysis; Phenotype; RNA, Ribosomal, 16S

The pattern of expression of the genes involved in the utilization of aryl beta-glucosides such as arbutin and salicin is different in the genus Shigella compared to Escherichia coli. The results presented here indicate that the homologue of the cryptic bgl operon of E. coli is conserved in Shigella sonnei and is the primary system involved in beta-glucoside utilization in the organism. The organization of the bgl genes in S. sonnei is similar to that of E. coli; however there are three major differences in terms of their pattern of expression. (i) The bglB gene, encoding phospho-beta-glucosidase B, is insertionally inactivated in S. sonnei. As a result, mutational activation of the silent bgl promoter confers an Arbutin-positive (Arb(+)) phenotype to the cells in a single step; however, acquiring a Salicin-positive (Sal(+)) phenotype requires the reversion or suppression of the bglB mutation in addition. (ii) Unlike in E. coli, a majority of the activating mutations (conferring the Arb(+) phenotype) map within the unlinked hns locus, whereas activation of the E. coli bgl operon under the same conditions is predominantly due to insertions within the bglR locus. (iii) Although the bgl promoter is silent in the wild-type strain of S. sonnei (as in the case of E. coli), transcriptional and functional analyses indicated a higher basal level of transcription of the downstream genes. This was correlated with a 1 bp deletion within the putative Rho-independent terminator present in the leader sequence preceding the homologue of the bglG gene. The possible evolutionary implications of these differences for the maintenance of the genes in the cryptic state are discussed.

Topics: Arbutin; Base Sequence; Benzyl Alcohols; DNA Primers; Escherichia coli; Evolution, Molecular; Gene Expression Regulation, Bacterial; Genes, Bacterial; Glucosides; Molecular Sequence Data; Mutation; Operon; Phenotype; Plasmids; Shigella sonnei; Species Specificity

The salAB genes of Azospirillum irakense KBC1, which encode two aryl-beta-glucosidases, are required for growth on salicin. In the 4-kb region upstream of the salAB genes, two additional genes, salC and salR, were identified. SalC shows characteristics of the outer membrane receptors in the FepA/FhuA family. The salC AB genes are transcribed as a polycistronic mRNA. The salR gene encodes a protein homologous to the LacI/GalR family of transcriptional repressors. Expression of the sal operon, measured by means of a salC-gusA translational fusion in A. irkense KBC1, requires the presence of aryl-beta-glucosides such as arbutin and salicin. Expression is markedly enhanced when a simple carbon source, like glucose, cellobiose or malate, is added to the medium. In a salR mutant, expression of the salC-gusA fusion does not require an aryl-beta-glucoside inducer. Expression of a salR-gusA fusion is constitutive. The product of arbutin hydrolysis (hydroquinone) partly inhibits the expression of a salC-gusA fusion in arbutin- or salicin-containing minimal medium. This effect is independent of SalR. Salicylalcohol, the hydrolysis product of salicin, also partly inhibits salC expression in a SalR-independent fashion, but only in salicin-containing minimal medium.

Topics: Amino Acid Sequence; Arbutin; Azospirillum; Bacterial Proteins; Benzyl Alcohols; beta-Glucosidase; Carrier Proteins; Cloning, Molecular; Enzyme Induction; Gene Expression Regulation, Bacterial; Genes, Bacterial; Glucosides; Hydrolysis; Hydroquinones; Molecular Sequence Data; Operon; Receptors, Cell Surface; Repressor Proteins; Sequence Analysis, DNA; Sequence Homology, Amino Acid; Transcription, Genetic

The rhizosphere nitrogen-fixing bacterium Azospirillum irakense KBC1 is able to grow on pectin and beta-glucosides such as cellobiose, arbutin, and salicin. Two adjacent genes, salA and salB, conferring beta-glucosidase activity to Escherichia coli, have been identified in a cosmid library of A. irakense DNA. The SalA and SalB enzymes preferentially hydrolyzed aryl beta-glucosides. A Delta(salA-salB) A. irakense mutant was not able to grow on salicin but could still utilize arbutin, cellobiose, and glucose for growth. This mutant could be complemented by either salA or salB, suggesting functional redundancy of these genes in salicin utilization. In contrast to this functional homology, the SalA and SalB proteins, members of family 3 of the glycosyl hydrolases, show a low degree of amino acid similarity. Unlike SalA, the SalB protein exhibits an atypical truncated C-terminal region. We propose that SalA and SalB are representatives of the AB and AB' subfamilies, respectively, in glycosyl hydrolase family 3. This is the first genetic implication of this beta-glucosidase family in the utilization of beta-glucosides for microbial growth.

Topics: Amino Acid Sequence; Arbutin; Azospirillum; Benzyl Alcohols; beta-Glucosidase; Cellobiose; Cloning, Molecular; Gene Deletion; Genetic Complementation Test; Glucose; Glucosides; Hydrogen-Ion Concentration; Isoelectric Point; Kinetics; Molecular Sequence Data; Phylogeny; Restriction Mapping; Sequence Homology, Amino Acid; Substrate Specificity; Temperature

A phosphotransferase-dependent aryl-beta-glucoside uptake and utilisation system (abg) was isolated from the ruminal Clostridium ("C. Longisporum"). The system is composed of three genes, abgG, abgF and abgA, and a number of regulatory regions, including terminator/antiterminator type stem-loop structures preceding the abgG and abgF genes. Similarity analysis of the proteins encoded by these genes indicated that they were responsible for the regulation of the abg system through antitermination (AbgG), the uptake and phosphorylation of aryl-beta-glucosides (AbgF) and the hydrolysis of the intracellular phosphorylated glycosides (AbgA). Experimental evidence for the functions of AbgF and AbgA was obtained. Although it was not possible to demonstrate any function for AbgG, a promoter 5' to the abgG gene was identified which was responsible for expression of the downstream genes. The abg system is remarkably similar to operons from the gram negative Enterobacteriaceae, both in the coding and non-coding regulatory regions.

Topics: Amino Acid Sequence; Animals; Arbutin; Bacterial Proteins; Base Sequence; Benzyl Alcohols; Biological Transport; Chymotrypsin; Cloning, Molecular; Clostridium; Gene Library; Genes, Bacterial; Glucosides; Glycoside Hydrolases; Membrane Transport Proteins; Molecular Sequence Data; Multienzyme Complexes; Phosphorylation; Rumen; Sequence Homology, Nucleic Acid

The expression of the putative operon bglPH of Bacillus subtilis was studied by using bglP'-lacZ transcriptional fusions. The bglP gene encodes an aryl-beta-glucoside-specific enzyme II of the phosphoenolpyruvate sugar:phosphotransferase system, whereas the bglH gene product functions as a phospho-beta-glucosidase. Expression of bglPH is regulated by at least two different mechanisms: (i) carbon catabolite repression and (ii) induction via an antitermination mechanism. Distinct deletions of the promoter region were created to determine cis-acting sites for regulation. An operatorlike structure partially overlapping the -35 box of the promoter of bglP appears to be the catabolite-responsive element of this operon. The motif is similar to that of amyO and shows no mismatches with respect to the consensus sequence established as the target of carbon catabolite repression in B. subtilis. Catabolite repression is abolished in both ccpA and ptsH1 mutants. The target of the induction by the substrate, salicin or arbutin, is a transcriptional terminator located downstream from the promoter of bglP. This structure is very similar to that of transcriptional terminators which regulate the induction of the B. subtilis sacB gene, the sacPA operon, and the Escherichia coli bgl operon. The licT gene product, a member of the BglG-SacY family of antitermination proteins, is essential for the induction process. Expression of bglP is under the negative control of its own gene product. The general proteins of the phosphoenolpyruvate-dependent phosphotransferase system are required for bglP expression. Furthermore, the region upstream from bglP, which reveals a high AT content, exerts a negative regulatory effect on bglP expression.

Topics: Arbutin; Bacillus subtilis; Bacterial Proteins; Base Sequence; Benzyl Alcohols; DNA-Binding Proteins; Enzyme Induction; Enzyme Repression; Gene Expression Regulation, Bacterial; Glucosidases; Glucosides; Molecular Sequence Data; Operator Regions, Genetic; Operon; Phosphoenolpyruvate Sugar Phosphotransferase System; Promoter Regions, Genetic; Recombinant Fusion Proteins; Repressor Proteins; Sequence Deletion; Terminator Regions, Genetic

The phytopathogenic bacterium Erwinia chrysanthemi, unlike other members of the family Enterobacteriaceae, is able to metabolize the beta-glucosides, arbutin, and salicin. A previous genetic analysis of the E. chrysanthemi arb genes, which mediate beta-glucoside metabolism, suggested that they were homologous to the Escherichia coli K-12 bgl genes. We have now determined the nucleotide sequence of a 5,065-bp DNA fragment containing three genes, arbG, arbF, and arbB. Deletion analysis, expression in minicell systems, and comparison with sequences of other proteins suggest that arbF and arbB encode a beta-glucoside-specific phosphotransferase system-dependent permease and a phospho-beta-glucosidase, respectively. The ArbF amino acid sequence shares 55% identity with that of the E. coli BglF permease and contains most residues thought to be important for a phosphotransferase. One change, however, was noted, since BglF Arg-625, presumably involved in phosphoryl transfer, was replaced by a Cys residue in ArbF. An analysis of the ArbB sequence led to the definition of a protein family which contained enzymes classified as phospho-beta-glucosidases, phospho-beta-galactosidases, beta-glucosidases, and beta-galactosidases and originating from gram-positive and gram-negative bacteria, archebacteria, and mammals, including humans. An analysis of this family allowed us (i) to speculate on the ways that these enzymes evolved, (ii) to identify a glutamate residue likely to be a key amino acid in the catalytic activity of each protein, and (iii) to predict that domain II of the human lactate-phlorizin hydrolase, which is involved in lactose intolerance, is catalytically nonactive. A comparison between the untranslated regions of the E. chrysanthemi arb cluster and the E. coli bgl operon revealed the conservation of two regions which, in the latter, are known to terminate transcription under noninducing conditions and be the target of the BglG transcriptional antiterminator under inducing conditions. ArbG was found to share a high level of similarity with the BglG antiterminator as well as with Bacillus subtilis SacT and SacY antiterminators, suggesting that ArbG functions as an antiterminator in regulating the expression of the E. chrysanthemi arb genes.

Topics: Amino Acid Sequence; Arbutin; Archaea; Base Sequence; Benzyl Alcohols; Biological Evolution; Cloning, Molecular; DNA Mutational Analysis; Erwinia; Escherichia coli; Glucosides; Glycoside Hydrolases; Lac Operon; Molecular Sequence Data; Multigene Family; Peptide Chain Termination, Translational; Pseudogenes; Recombinant Fusion Proteins; Regulatory Sequences, Nucleic Acid; Sequence Homology, Nucleic Acid

Topics: Arbutin; Benzyl Alcohols; Cellobiose; Escherichia coli; Genes, Bacterial; Glucosides; Operon; Restriction Mapping

We have analyzed the functions encoded by the bgl operon in Escherichia coli K-12. Based on the ability of cloned regions of the operon to complement a series of Bgl- point mutations, we show that the three bgl structural genes, bglC, bglS, and bglB, are located downstream of the regulatory locus bglR in the order indicated. Using a bgl-lacZ transcriptional fusion, we show that bglC and bglS are involved in regulating operon expression. The presence of the bglC gene in trans is absolutely required for the expression of the fusion, which is constitutive when only the bglC gene is present. When the bglC and the bglS genes are both present in the cell, expression of the fusion requires a beta-glucoside inducer. From these observations, we conclude that (i) the bglC gene encodes a positive regulatory of bgl operon expression and (ii) the bglS gene encodes a negative regulator of operon expression, causing the requirement for a beta-glucoside inducer. These conclusions are supported by our observations that (i) a majority of bglC mutants exhibits a Bgl- phenotype, whereas rare trans-dominant mutations in bglC result in constitutive expression of the bgl operon and the fusion, and (ii) mutations in the bglS gene lead to constitutive expression of the fusion. Based on several lines of evidence presented, we propose that the bglS gene product has an additional role as a component of the beta-glucoside transport system.

Topics: Arbutin; Benzyl Alcohols; Chromosome Mapping; Escherichia coli; Gene Expression Regulation; Genes; Genes, Bacterial; Genes, Regulator; Glucosides; Glycosides; Mutation; Operon; Phosphoenolpyruvate Sugar Phosphotransferase System; Promoter Regions, Genetic; Recombinant Fusion Proteins; Transcription, Genetic

The cellobiose catabolic system of Escherichia coli K12 is being used to study the role of cryptic genes in evolution of new functions. Escherichia coli does not use beta-glucoside sugars; however, mutations in several loci can activate the cryptic bgl operon and permit growth on the beta-glucoside sugars arbutin and salicin. Such Bgl+ mutants do not use cellobiose, which is the most common beta-glucoside in nature. We have isolated a Cel+ (cellobiose-utilizing) mutant from a Bgl+ mutant of E. coli K12. The Cel+ mutant grows well on cellobiose, arbutin, and salicin. Genes for utilization of these beta-glucosides are located at 37.8 min on the E. coli map. The genes of the bgl operon are not involved in cellobiose utilization. Introduction of a deletion covering bgl does not affect the ability to utilize cellobiose, arbutin, or salicin, indicating that the new Cel+ genes provide all three functions. Spontaneous cellobiose negative mutants also become arbutin and salicin negative. Analysis of beta-glucoside positive revertants of these mutants indicates that there are separate loci for utilization of each of the beta-glucoside sugars. The genes are closely linked and may be activated from a single locus. A fourth gene at an unknown location increases the growth rate on cellobiose. The cel genes constitute a second cryptic system for beta-glucoside utilization in E. coli K12.

Topics: Arbutin; Benzyl Alcohols; Biological Evolution; Cellobiose; Disaccharides; Escherichia coli; Genes, Bacterial; Glucosides; Mutation

A study of the abilities of 23 strains of Pseudomonas maltophilia to hydrolyze synthetically-prepared and naturally-occurring glycosides is presented. Direct detection of liberated aglycones was used to determine hydrolysis of the five most-commonly-used glycosides (amygdalin, arbutin, esculin, ONPG (o-nitrophenyl-beta-d-galactoside), and salicin). The capabilities of the strain for acid production from 17 glycoside substrates were also determined using a medium designed to minimize the production of acid-neutralizing end-products from peptones.

Topics: Amygdalin; Arbutin; Benzyl Alcohols; Esculin; Glucosides; Glycosides; Methods; Nitrophenylgalactosides; Pseudomonas