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Heterotrophic microbes rely on host‐derived carbon sources for their growth and survival. Depriving pathogens of plant carbon is therefore a promising strategy for protecting plants from disease and reducing yield losses. Importantly, this carbon starvation‐mediated resistance is expected to be more broad‐spectrum and durable than race‐specific R‐gene‐mediated resistance. Although sugars are well characterized as major carbon sources for bacteria, emerging evidence suggests that plant‐derived lipids are likely to be an essential carbon source for some fungal microbes, particularly biotrophs. Here, we comprehensively discuss the dual roles of carbon sources (mainly sugars and lipids) and their transport processes in immune signalling and microbial nutrition. We summarize recent findings revealing the crucial roles of lipids as susceptibility factors at all stages of pathogen infection. In particular, we discuss the potential pathways by which lipids and other plant carbon sources are delivered to biotrophs, including protein‐mediated transport, vesicle trafficking and autophagy. Finally, we highlight knowledge gaps and offer suggestions for clarifying the mechanisms that underlie nutrient uptake by biotrophs, providing guidance for future research on the application of carbon starvation–mediated resistance. Summary statement: This review summarizes the roles of host carbon sources in plant–microbe interactions, highlights the carbon nutrition and susceptibility factor roles of host lipids for biotrophic fungi and outlines several host transport processes that potentially contribute to fungal carbon acquisition. [ABSTRACT FROM AUTHOR]

Summary: The mutualistic interaction between ectomycorrhizal fungi and trees is characterized by the coordinated exchange of soil nutrients with soluble sugars. Despite the importance of this process, the precise mechanism by which sugars are transported from host roots to colonizing hyphae remains unclear.This study aimed to identify the specific membrane transporters responsible for the unloading of sugars at the symbiotic interface, with a focus on the role of the root Sugars Will Eventually Be Exported Transporter (SWEET) uniporters.Our study used RNA sequencing and quantitative PCR to identify PtaSWEET gene expression in Populus tremula × alba–Laccaria bicolor ectomycorrhizal root tips. Our results suggest that symbiosis‐induced PtaSWEET1c is primarily responsible for transporting glucose and sucrose, as demonstrated by the yeast assays. Moreover, we used a promoter‐YFP reporter to confirm the localization of the PtaSWEET1c expression in cortical cells of ectomycorrhizal rootlets, supporting its major role in supplying glucose at the symbiotic interface. Furthermore, our observations confirmed the localization of PtaSWEET1c‐GFP in the plasma membrane. The inactivation of PtaSWEET1c reduced ectomycorrhizal root formation and 13C translocation to ectomycorrhizal roots.Our findings highlight the crucial role of PtaSWEET1c in facilitating glucose and sucrose transport at the symbiotic interface of Populus–L. bicolor symbiosis. [ABSTRACT FROM AUTHOR]

Summary How carbon (sucrose) and nitrogen (amino acid) accumulation is coordinatively controlled in cereal grains remains largely enigmatic. We found that overexpression of the strigolactone (SL) biosynthesis gene CAROTENOID CLEAVAGE DIOXYGENASE 8 (CCD8) resulted in greater ear diameter and enhanced sucrose and amino acid accumulation in maize kernels. Loss of ZmCCD8 function reduced kernel growth with lower sugar and amino acid concentrations. Transcriptomic analysis showed down‐regulation of the transcription factors ZmMYB42 and ZmMYB63 in ZmCCD8 overexpression alleles and up‐regulation in zmccd8 null alleles. Importantly, ZmMYB42 and ZmMYB63 were negatively regulated by the SL signalling component UNBRANCHED 3, and repressed expression of the sucrose transporters ZmSWEET10 and ZmSWEET13c and the lysine/histidine transporter ZmLHT14. Consequently, null alleles of ZmMYB42 or ZmMYB63 promoted accumulation of soluble sugars and free amino acids in maize kernels, whereas ZmLHT14 overexpression enhanced amino acid accumulation in kernels. Moreover, overexpression of the SL receptor DWARF 14B resulted in more sucrose and amino acid accumulation in kernels, down‐regulation of ZmMYB42 and ZmMYB63 expression, and up‐regulation of ZmSWEETs and ZmLHT14 transcription. Together, we uncover a distinct SL signalling pathway that regulates sucrose and amino acid accumulation in kernels. Significant association of two SNPs in the 5′ upstream region of ZmCCD8 with ear and cob diameter implicates its potential in breeding toward higher yield and nitrogen efficiency. [ABSTRACT FROM AUTHOR]

Summary: In photosynthetic cells, plants convert carbon dioxide to sugars that can be moved between cellular compartments by transporters before being subsequently metabolized to support plant growth and development. Most pathogens cannot synthesize sugars directly but have evolved mechanisms to obtain plant‐derived sugars as C resource for successful infection and colonization. The availability of sugars to pathogens can determine resistance or susceptibility. Here, we summarize current progress on the roles of sugar transporters in plant–pathogen interactions. We highlight how transporters are manipulated antagonistically by both host and pathogens in competing for sugars. We examine the potential application of this target in resistance breeding and discuss opportunities and challenges for the future. [ABSTRACT FROM AUTHOR]

Beneficial Bacillus subtilis (BS) symbiosis could combat root pathogenesis, but it relies on root‐secreted sugars. Understanding the molecular control of sugar flux during colonization would benefit biocontrol applications. The SWEET (Sugar Will Eventually Be Exported Transporter) uniporter regulates microbe‐induced sugar secretion from roots; thus, its homologs may modulate sugar distribution upon BS colonization. Quantitative polymerase chain reaction revealed that gene transcripts of SWEET2, but not SWEET16 and 17, were significantly induced in seedling roots after 12 h of BS inoculation. Particularly, SWEET2‐β‐glucuronidase fusion proteins accumulated in the apical mature zone where BS abundantly colonized. Yet, enhanced BS colonization in sweet2 mutant roots suggested a specific role for SWEET2 to constrain BS propagation, probably by limiting hexose secretion. By employing yeast one‐hybrid screening and ectopic expression in Arabidopsis protoplasts, the transcription factor AHL29 was identified to function as a repressor of SWEET2 expression through the AT‐hook motif. Repression occurred despite immunity signals. Additionally, enhanced SWEET2 expression and reduced colonies were specifically detected in roots of BS‐colonized ahl29 mutant. Taken together, we propose that BS colonization may activate repression of AHL29 on SWEET2 transcription that would be enhanced by immunity signals, thereby maintaining adequate sugar secretion for a beneficial Bacillus association. Summary statement: Arabidopsis vacuolar SWEET2 transporter limits root sugar secretion upon colonization with beneficial Bacillus subtilis. The AT‐hook protein AHL29 binds to the SWEET2 promoter to repress its immunity‐induced expression, thereby promoting adequate carbon for a mutualistic association with Bacillus. [ABSTRACT FROM AUTHOR]

Cell wall polysaccharides play key roles in fungal development, virulence, and resistance to the plant immune system, and are synthesized from many nucleotide sugars in the endoplasmic reticulum (ER)‐Golgi secretory system. Nucleotide sugar transporters (NSTs) are responsible for transporting cytosolic‐derived nucleotide sugars to the ER lumen for processing, but their roles in plant‐pathogenic fungi remain to be revealed. Here, we identified two important NSTs, NST1 and NST2, in the rice blast fungus Magnaporthe oryzae. Both NSTs were localized in the ER, which was consistent with a function in transporting nucleotide sugar for processing in the ER. Sugar transport property analysis suggested that NST1 is involved in transportation of mannose and glucose, while NST2 is only responsible for mannose transportation. Accordingly, deletion of NSTs resulted in a significant decrease in corresponding soluble saccharides abundance and defect in sugar utilization. Moreover, both NSTs played important roles in cell wall integrity, were involved in asexual development, and were required for full virulence. The NST mutants exhibited decreasing external glycoproteins and exposure of inner chitin, which resulted in activation of the host defence response. Altogether, our results revealed that two sugar transporters are required for fungal cell wall polysaccharides accumulation and full virulence of M. oryzae. [ABSTRACT FROM AUTHOR]

Summary: More than half of the world's food is provided by cereals, as humans obtain >60% of daily calories from grains. Producing more carbohydrates is always the final target of crop cultivation. The carbohydrate partitioning pathway directly affects grain yield, but the molecular mechanisms and biological functions are poorly understood, including rice (Oryza sativa L.), one of the most important food sources. Here, we reported a prolonged grain filling duration mutant 1 (gfd1), exhibiting a long grain‐filling duration, less grain number per panicle and bigger grain size without changing grain weight. Map‐based cloning and molecular biological analyses revealed that GFD1 encoded a MATE transporter and expressed high in vascular tissues of the stem, spikelet hulls and rachilla, but low in the leaf, controlling carbohydrate partitioning in the stem and grain but not in the leaf. GFD1 protein was partially localized on the plasma membrane and in the Golgi apparatus, and was finally verified to interact with two sugar transporters, OsSWEET4 and OsSUT2. Genetic analyses showed that GFD1 might control grain‐filling duration through OsSWEET4, adjust grain size with OsSUT2 and synergistically modulate grain number per panicle with both OsSUT2 and OsSWEET4. Together, our work proved that the three transporters, which are all initially classified in the major facilitator superfamily family, could control starch storage in both the primary sink (grain) and temporary sink (stem), and affect carbohydrate partitioning in the whole plant through physical interaction, giving a new vision of sugar transporter interactome and providing a tool for rice yield improvement. [ABSTRACT FROM AUTHOR]

SUMMARY: Developing seed depends on sugar supply for its growth and yield formation. Maize (Zea mays L.) produces the largest grains among cereals. However, there is a lack of holistic understanding of the transcriptional landscape of genes controlling sucrose transport to, and utilization within, maize grains. By performing in‐depth data mining of spatio‐temporal transcriptomes coupled with histological and heterologous functional analyses, we identified transporter genes specifically expressed in the maternal–filial interface, including (i) ZmSWEET11/13b in the placento‐chalazal zone, where sucrose is exported into the apoplasmic space, and (ii) ZmSTP3, ZmSWEET3a/4c (monosaccharide transporters), ZmSUT1, and ZmSWEET11/13a (sucrose transporters) in the basal endosperm transfer cells for retrieval of apoplasmic sucrose or hexoses after hydrolysis by extracellular invertase. In the embryo and its surrounding regions, an embryo‐localized ZmSUT4 and a cohort of ZmSWEETs were specifically expressed. Interestingly, drought repressed those ZmSWEETs likely exporting sucrose but enhanced the expression of most transporter genes for uptake of apoplasmic sugars. Importantly, this drought‐induced fluctuation in gene expression was largely attenuated by an increased C supply via controlled pollination, indicating that the altered gene expression is conditioned by C availability. Based on the analyses above, we proposed a holistic model on the spatio‐temporal expression of genes that likely govern sugar transport and utilization across maize maternal and endosperm and embryo tissues during the critical stage of grain set. Collectively, the findings represent an advancement towards a holistic understanding of the transcriptional landscape underlying post‐phloem sugar transport in maize grain and indicate that the drought‐induced changes in gene expression are attributable to low C status. Significance Statement: Although maize (Zea mays L.) produces the largest grain among cereals through a small 'gateway' for assimilates, indicating a strong sink capacity, there is a lack of holistic understanding of the molecular basis underlying sugar import into the grains. Here, we uncovered a cohort of key genes controlling sugar delivery to the endosperm and embryo and further found that drought repressed those genes exporting sucrose into the apoplasmic space but enhanced those genes involved in uptake of sugars by the filial tissues. [ABSTRACT FROM AUTHOR]

Sugar transporters (STs), which mainly mediate cellular sugar exchanges, play critical physiological roles in living organisms, and they may be responsible for sugar exchanges among various insect tissues. However, the molecular and physiological functions of insect STs are largely unknown. Here, 16 STs of Helicoverpa armigera were identified. A phylogenetic analysis classified the putative HaSTs into 12 sub‐families, and those identified in this study were distributed into 6 sub‐families. Real‐time polymerase chain reaction indicated that the 16 HaSTs had diverse tissue‐specific expression levels. One transporter, HaST10, was highly expressed in thoracic muscles. A functional study using a Xenopus oocyte expression system revealed that HaST10 mediated both H+‐driven trehalose and Na+‐driven glucose antiport activities with high transport efficiency and low affinity levels. A HaST10 knockout clearly impaired the performance of H. armigera. Thus, HaST10 may participate in sugar‐supply regulation and have essential physiological roles in H. armigera. [ABSTRACT FROM AUTHOR]

Summary: Abiotic stresses affect plant growth and development by causing cellular damage and/or restricting resources. Plants often respond to stresses through abscisic acid (ABA) signaling. Exogenous ABA application can therefore be used to mimic stress responses, which can be overridden by glucose (Glc) addition during seed germination. It remains unclear whether ABA‐mediated germination inhibition is due to regional or global suppression of Glc availability in germinating Arabidopsis seeds.We used a genetically engineered Förster resonance energy transfer (FRET) sensor to ascertain whether ABA affects the spatiotemporal distribution of Glc, 14C‐Glc uptake assays to track potential effects of ABA on sugar import, and transcriptome and mutant analyses to identify genes associated with Glc availability that are involved in ABA‐inhibited seed germination.Abscisic acid limits Glc in the hypocotyl largely by suppressing sugar allocation as well as altering sugar metabolism. Mutant plants carrying loss‐of‐function ABA‐inducible sucrose‐phosphate synthase (SPS) genes accumulated more Glc, leading to ABA‐insensitive germination. We reveal that Glc antagonizes ABA by globally counteracting the ABA influence at the transcript level, including expansin (EXP) family genes suppressed by ABA.This study presents a new perspective on how ABA affects Glc distribution, which likely reflects what occurs when seeds are subjected to abiotic stresses such as drought and salt stress. [ABSTRACT FROM AUTHOR]

SUMMARY: The sugar transporter (ST) family is considered to be the most important gene family for sugar accumulation, but limited information about the ST family in the important sugar‐yielding crop Saccharum is available due to its complex genetic background. Here, 105 ST genes were identified and clustered into eight subfamilies in Saccharum spontaneum. Comparative genomics revealed that tandem duplication events contributed to ST gene expansions of two subfamilies, PLT and STP, in S. spontaneum, indicating an early evolutionary step towards high sugar content in Saccharum. The analyses of expression patterns were based on four large datasets with a total of 226 RNA sequencing samples from S. spontaneum and Saccharum officinarum. The results clearly demonstrated 50 ST genes had different spatiotemporal expression patterns in leaf tissues, 10 STs were specifically expressed in the stem, and 10 STs responded to the diurnal rhythm. Heterologous expression experiments in the defective yeast strain EBY.VW4000 indicated STP13, pGlcT2, VGT3, and TMT4 are the STs with most affinity for glucose/fructose and SUT1_T1 has the highest affinity to sucrose. Furthermore, metabolomics analysis suggested STP7 is a sugar starvation‐induced gene and STP13 has a function in retrieving sugar in senescent tissues. PLT11, PLT11_T1, TMT3, and TMT4 contributed to breaking the limitations of the storage sink. SUT1, SUT1_T1, PLT11, TMT4, pGlcT2, and VGT3 responded for different functions in these two Saccharum species. This study demonstrated the evolutionary expansion and functional divergence of the ST gene family and will enable the further investigation of the molecular mechanism of sugar metabolism in Saccharum. Significance Statement: An early evolutionary step towards high sugar content in Saccharum was indicated to be caused by recent tandem duplication events in Saccharum spontaneum, and functional divergence of the ST family between S. spontaneum and Saccharum officinarum was further unveiled based on heterologous expression experiments, a large set of transcriptome data, and metabolomics analysis. This study will enable the further investigation of the molecular mechanism of sugar metabolism in Saccharum. [ABSTRACT FROM AUTHOR]

Summary: Unloading sugar from sink phloem by transporters is complex and much remains to be understood about this phenomenon in the watermelon fruit.Here, we report a novel vacuolar sugar transporter (ClVST1) identified through map‐based cloning and association study, whose expression in fruit phloem is associated with accumulation of sucrose (Suc) in watermelon fruit. ClVST197 knockout lines show decreased sugar content and total biomass, whereas overexpression of ClVST197 increases Suc content.Population genomic and subcellular localization analyses strongly suggest a single‐base change at the coding region of ClVST197 as a major molecular event during watermelon domestication, which results in the truncation of 45 amino acids and shifts the localization of ClVST197 to plasma membranes in sweet watermelons. Molecular, biochemical and phenotypic analyses indicate that ClVST197 is a novel sugar transporter for Suc and glucose efflux and unloading.Functional characterization of ClVST1 provides a novel strategy to increase sugar sink potency during watermelon domestication. [ABSTRACT FROM AUTHOR]

Keywords: fibre initiation; fibre elongation; secondary cell wall deposition; sucrose; sugar transporter; reactive oxygen species EN fibre initiation fibre elongation secondary cell wall deposition sucrose sugar transporter reactive oxygen species 1092 1094 3 06/15/21 20210601 NES 210601 Cotton fibre is a single cell that derived from the ovule epidermis. We also suggest a strategy to improve cotton quality and yield through sugar manipulation: up-regulating the sugar content in fibre initiation and early elongation stage to increase fibre number (thus enhancing fibre yield), while down-regulating the sugar content in fibre SCW synthesis period to increase fibre length and fibre strength. [Extracted from the article]

Fumonisin B1 (FB1), a polyketide mycotoxin produced by Fusarium verticillioides during the colonization of maize kernels, is detrimental to human and animal health. FST1 encodes a putative protein with 12 transmembrane domains; however, its function remains unknown. The FST1 gene is highly expressed by the fungus in the endosperm of maize kernels compared with the levels of expression in germ tissues. Previous research has shown that FST1 affects FB1 production, virulence, hydrogen peroxide resistance, hydrophobicity and macroconidia production. Here, we examine the phylogeny of FST1, its expression in a Saccharomyces cerevisiae strain lacking a functional myo-inositol transporter (ITR1) and the effect of amino acid changes in the central loop and C-terminus regions of FST1 on functionality. The results indicate that expression of FST1 in an ITR1 mutant strain restores growth on myo-inositol medium to wild-type levels and restores the inhibitory effects of FB1, suggesting that FST1 can transport both myo-inositol and FB1 into yeast cells. Our results with engineered FST1 also indicate that amino acids in the central loop and C-terminus regions are important for FST1 functionality in both S. cerevisiae and F. verticillioides. Overall, this research has established the first characterized inositol transporter in filamentous fungi and has advanced our knowledge about the global regulatory functions of FST1. [ABSTRACT FROM AUTHOR]

ABSTRACT Efficient glucose-xylose co-utilization is critical for economical biofuel production from lignocellulosic biomass. To enable glucose-xylose co-utilization, a highly active xylose specific transporter without glucose inhibition is desirable. However, our understanding of the structure-activity/specificity relationship of sugar transporters in general is limited, which hinders our ability to engineer xylose-specific transporters. In this study, via homology modeling and analysis of hexose sugar transporter HXT14 mutants, we identified a highly conserved YYX(T/P) motif that plays an important role in controlling the activity and specificity of sugar transporters. We demonstrated that mutating the two tyrosine residues of the motif to phenylalanine, respectively, improved glucose transport capacity across several different sugar transporters. Furthermore, we illustrated that by engineering the fourth position in the YYX(T/P) motif, the sugar specificity of transporters was significantly altered or even reversed towards xylose. Finally, using the engineered sugar transporter, genuine glucose-xylose co-fermentation was achieved. Biotechnol. Bioeng. 2016;113: 1460-1467. © 2016 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

The transport of sugars across the plasma membrane is a critical step in the utilization of glucose and fructose by Saccharomyces cerevisiae during must fermentations. Variations in the molecular structure of hexose transporters and kinases may affect the ability of wine yeast strains to finish sugar fermentation, even under stressful wine conditions. In this context, we sequenced and compared genes encoding the hexose transporter Hxt3p and the kinases Hxk1p/Hxk2p of Saccharomyces strains and interspecies hybrids with different industrial usages and regional backgrounds. The Hxt3p primary structure varied in a small set of amino acids, which characterized robust yeast strains used for the production of sparkling wine or to restart stuck fermentations. In addition, interspecies hybrid strains, previously isolated at the end of spontaneous fermentations, revealed a common amino acid signature. The location and potential influence of the amino acids exchanges is discussed by means of a first modelled Hxt3p structure. In comparison, hexokinase genes were more conserved in different Saccharomyces strains and hybrids. Thus, molecular variants of the hexose carrier Hxt3p, but not of kinases, correlate with different fermentation performances of yeast. Copyright © 2015 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Plant sugar will eventually be exported transporter (SWEET) sugar transporters have been implicated in various developmental processes where sugar efflux is essential, including sucrose loading of phloem for long-distance sugar transport, nectar secretion, embryo and pollen nutrition, and maintenance of sugar homeostasis in plant organs. Notably, these transporters are selectively targeted by pathogens to gain access to host sugars. In most cases, when SWEET function is blocked, the growth and virulence of the pathogen is also reduced. There is growing evidence to suggest that the lifestyle of the pathogen may dictate which SWEET or set of SWEET genes are recruited for pathogen growth and proliferation. Furthermore, SWEET transporters may also play a role in abiotic stress tolerance by enabling plant growth under unfavorable environmental conditions. This review provides an overview of the diverse functions of SWEET proteins in plant development, pathogen nutrition, and abiotic stress tolerance. In addition, utility of the model legume Medicago truncatula as a tool to elucidate SWEET function in diverse host-microbe interactions is discussed. © 2015 IUBMB Life, 67(7):461-471, 2015 [ABSTRACT FROM AUTHOR]

The antipsychotic drug olanzapine is widely prescribed to treat schizophrenia and other psychotic disorders. However, it often causes unwanted side effects, including diabetes, due to disruption of insulin-dependant glucose metabolism through a mechanism yet to be elucidated. To determine if olanzapine can affect the first step in glucose metabolism – glucose transport inside cells – we investigated the effect of this drug on the transport activity of a model glucose transporter. The glucose transporter from Staphylococcus epidermidis (GlcP Se ) is specific for glucose, inhibited by various human glucose transporter (GLUT) inhibitors, has high sequence and structure homology to GLUTs, and is readily amenable to transport assay, mutagenesis, and computational modeling. We found that olanzapine inhibits glucose transport of GlcP Se with an IC 50 0.9 ± 0.1 mM. Computational docking of olanzapine to the GlcP Se structure revealed potential binding sites that were further examined through mutagenesis and transport assay to identify residues important for olanzapine inhibition. These investigations suggest that olanzapine binds in a polar region of the cytosolic part of the transporter, and interacts with residues R129, strictly conserved in all GLUTs, and N136, conserved in only a few GLUTs, including the insulin-responsive GLUT4. We propose that olanzapine inhibits GlcP Se by impeding the alternating opening and closing of the substrate cavity necessary for glucose transport. It accomplishes this by disrupting a key salt bridge formed by conserved residues R129 and E362, that stabilizes the outward-facing conformation of the transporter. [ABSTRACT FROM AUTHOR]

•In Arabidopsis, the compartmentation of sugars into vacuoles is known to be facilitated by sugar transporters. However, vacuolar sugar transporters have not been studied in detail in other plant species. •To characterize the rice ( Oryza sativa) tonoplast monosaccharide transporters, OsTMT1 and OsTMT2, we analysed their subcellular localization using green fluorescent protein (GFP) and expression patterns using reverse-transcription polymerase chain reaction (RT-PCR), performed histochemical β-glucuronidase (GUS) assay and in situ hybridization analysis, and assessed sugar transport ability using isolated vacuoles. •Expression of OsTMT–GFP fusion protein in rice and Arabidopsis revealed that the OsTMTs localize at the tonoplast. Analyses of OsTMT promoter- GUS transgenic rice indicated that OsTMT1 and OsTMT2 are highly expressed in bundle sheath cells, and in vascular parenchyma and companion cells in leaves, respectively. Both genes were found to be preferentially expressed in the vascular tissues of roots, the palea/lemma of spikelets, and in the main vascular tissues and nucellar projections on the dorsal side of the seed coats. Glucose uptake studies using vacuoles isolated from transgenic mutant Arabidopsis ( tmt1-2-3) expressing OsTMT1 demonstrated that OsTMTs are capable of transporting glucose into vacuoles. •Based on expression analysis and functional characterization, our present findings suggest that the OsTMTs play a role in vacuolar glucose storage in rice. [ABSTRACT FROM AUTHOR]

The crystal structures of Thermoactinomyces  vulgaris cyclo/maltodextrin-binding protein ( TvuCMBP) complexed with α-cyclodextrin (α-CD), β-cyclodextrin (β-CD) and maltotetraose (G4) have been determined. A common functional conformational change among all solute-binding proteins involves switching from an open form to a closed form, which facilitates transporter binding. Escherichia  coli maltodextrin-binding protein ( EcoMBP), which is structurally homologous to TvuCMBP, has been determined to adopt the open form when complexed with β-CD and the closed form when bound to G4. Here, we show that, unlike EcoMBP, TvuCMBP–α-CD and TvuCMBP–β-CD adopt the closed form when complexed, whereas TvuCMBP–G4 adopts the open form. Only two glucose residues are evident in the TvuCMBP–G4 structure, and these bind to the C-domain of TvuCMBP in a manner similar to the way in which maltose binds to the C-domain of EcoMBP. The superposition of TvuCMBP–α-CD, TvuCMBP–β-CD and TvuCMBP–γ-CD shows that the positions and the orientations of three glucose residues in the cyclodextrin molecules overlay remarkably well. In addition, most of the amino acid residues interacting with these three glucose residues also participate in interactions with the two glucose residues in TvuCMBP–G4, regardless of whether the protein is in the closed or open form. Our results suggest that the mechanisms by which TvuCMBP changes from the open to the closed conformation and maintains the closed form appear to be different from those of EcoMBP, despite the fact that the amino acid residues responsible for the initial binding of the ligands are well conserved between TvuCMBP and EcoMBP. [ABSTRACT FROM AUTHOR]

The crystal structure of a Thermoactinomyces vulgaris cyclo/maltodextrin-binding protein ( TvuCMBP) complexed with γ-cyclodextrin has been determined. Like Escherichia coli maltodextrin-binding protein ( EcoMBP) and other bacterial sugar-binding proteins, TvuCMBP consists of two domains, an N- and a C-domain, both of which are composed of a central β-sheet surrounded by α-helices; the domains are joined by a hinge region containing three segments. γ-Cyclodextrin is located at a cleft formed by the two domains. A common functional conformational change has been reported in this protein family, which involves switching from an open form to a sugar-transporter bindable form, designated a closed form. The TvuCMBP–γ-cyclodextrin complex structurally resembles the closed form of EcoMBP, indicating that TvuCMBP complexed with γ-cyclodextrin adopts the closed form. The fluorescence measurements also showed that the affinities of TvuCMBP for cyclodextrins were almost equal to those for maltooligosaccharides. Despite having similar folds, the sugar-binding site of the N-domain part of TvuCMBP and other bacterial sugar-binding proteins are strikingly different. In TvuCMBP, the side-chain of Leu59 protrudes from the N-domain part into the sugar-binding cleft and orients toward the central cavity of γ-cyclodextrin, thus Leu59 appears to play the key role in binding. The cleft of the sugar-binding site of TvuCMBP is also wider than that of EcoMBP. These findings suggest that the sugar-binding site of the N-domain part and the wide cleft are critical in determining the specificity of TvuCMBP for γ-cyclodextrin. [ABSTRACT FROM AUTHOR]