Evi1 upregulates Fbp1 and supports progression of acute myeloid leukemia through pentose phosphate pathway activation.

Evi1 is a transcription factor essential for the development as well as progression of acute myeloid leukemia (AML) and high Evi1 AML is associated with extremely poor clinical outcome. Since targeting metabolic vulnerability is the emerging therapeutic strategy of cancer, we herein investigated a novel therapeutic target of Evi1 by analyzing transcriptomic, epigenetic, and metabolomic profiling of mouse high Evi1 leukemia cells. We revealed that Evi1 overexpression and Evi1-driven leukemic transformation upregulate transcription of gluconeogenesis enzyme Fbp1 and other pentose phosphate enzymes with interaction between Evi1 and the enhancer region of these genes. Metabolome analysis using Evi1-overexpressing leukemia cells uncovered pentose phosphate pathway upregulation by Evi1 overexpression. Suppression of Fbp1 as well as pentose phosphate pathway enzymes by shRNA-mediated knockdown selectively decreased Evi1-driven leukemogenesis in vitro. Moreover, pharmacological or shRNA-mediated Fbp1 inhibition in secondarily transplanted Evi1-overexpressing leukemia mouse significantly decreased leukemia cell burden. Collectively, targeting FBP1 is a promising therapeutic strategy of high Evi1 AML.

miR-18a-5p Targets FBP1 to Promote Proliferation, Migration, and Invasion of Liver Cancer Cells and Inhibit Cell Apoptosis.

Liver cancer is one of the most aggressive malignant tumors. It is significant to understand the molecular mechanism of liver cancer cells to develop new treatment plans. Studies have identified that FBP1 serves as a cancer inhibitor gene. To research the effect mechanism of FBP1 in liver cancer cells, bioinformatics analysis was performed to study its expression in liver cancer tissue. Survival analysis was also performed. Moreover, starBase database was applied to predict upstream regulatory genes of FBP1. Dual-luciferase assay was performed to testify their targeted relationship. The mRNA and protein expression levels of FBP1 in liver cancer cells were detected by qRT-PCR and western blot, respectively. Cell viability was analyzed by CCK-8 assay. The migratory and invasive abilities of cells were analyzed by Transwell assay. The apoptosis of liver cancer cells was detected by flow cytometry. The results showed that the expression of FBP1 was downregulated in liver cancer tissue and cells. FBP1 low expression was correlated with the poor prognosis of patients. miR-18a-5p could inhibit FBP1 expression. Overexpression of FBP1 could inhibit the progression of liver cancer cells and promote cell apoptosis. Overexpressing miR-18a-5p could promote the progression of liver cancer cells and inhibit cell apoptosis. However, overexpressing FBP1 simultaneously could reverse the effect. miR-18a-5p and FBP1 are expected to be candidates for liver cancer treatment.

Quantitative determination of Cpd118, A novel FBPase inhibitor, in dog plasma by HPLC-MS/MS.

Aim: A HPLC-MS/MS method was first developed and validated for the quantification of Cpd118, a novel fructose-1, 6-bisphosphatase inhibitor for controlling gluconeogenesis in Type 2 diabetes mellitus. Materials & methods: Cpd118 was extracted from dog plasma following acetonitrile protein precipitation, separated by HPLC on a CAPCELL PAK ADME column (3.5 µm, 2.1 mm × 100 mm) and quantified using negative heated electrospray ion source-MS/MS. Results: Cpd118 was quantified from plasma using the method described above over a linear range of 10-20,000 ng/ml, with interday and intraday assay accuracy from -11.78 to 4.01% and the precision was ≤11.15%. Conclusion: The method was sensitive and selective for the quantification of Cpd118 and was successfully used to the pharmacokinetic and bioavailability study of Cpd118 in dogs.

Discovery of N-Arylsulfonyl-Indole-2-Carboxamide Derivatives as Potent, Selective, and Orally Bioavailable Fructose-1,6-Bisphosphatase Inhibitors-Design, Synthesis, In Vivo Glucose Lowering Effects, and X-ray Crystal Complex Analysis.

Liver fructose-1,6-bisphosphatase (FBPase) is a key enzyme in the gluconeogenesis pathway. Inhibiting FBPase activity represents a potential treatment for type 2 diabetes mellitus. A series of novel N-arylsulfonyl-4-arylamino-indole-2-carboxamide derivatives have been disclosed as FBPase inhibitors. Through extensive structure-activity relationship investigations, a promising candidate molecule Cpd118 [sodium (7-chloro-4-((3-methoxyphenyl)amino)-1-methyl-1H-indole-2-carbonyl] [(4-methoxyphenyl)sulfonyl)amide] has been identified with high inhibitory activity against human liver FBPase (IC50, 0.029 ± 0.006 µM) and high selectivity relative to the other six AMP-binding enzymes. Importantly, Cpd118 produced significant glucose-lowering effects on both type 2 diabetic KKAy mice and ZDF rats as demonstrated by substantial reductions in the fasting and postprandial blood glucose levels, as well as the HbA1c level. Furthermore, Cpd118 elicited a favorable pharmacokinetic profile with an oral bioavailability of 99.1%. Moreover, the X-ray crystal structure of the Cpd118-FBPase complex was resolved, which revealed a unique binding mode and provided a structural basis for its high potency and selectivity.

Biological evaluation and SAR analysis of novel covalent inhibitors against fructose-1,6-bisphosphatase.

Fructose-1,6-bisphosphatase (FBPase) is an attractive target for affecting the GNG pathway. In our previous study, the C128 site of FBPase has been identified as a new allosteric site, where several nitrovinyl compounds can bind to inhibit FBPase activity. Herein, a series of nitrostyrene derivatives were further synthesized, and their inhibitory activities against FBPase were investigated in vitro. Most of the prepared nitrostyrene compounds exhibit potent FBPase inhibition (IC50 < 10 µM). Specifically, when the substituents of F, Cl, OCH3, CF3, OH, COOH, or 2-nitrovinyl were installed at the R2 (meta-) position of the benzene ring, the FBPase inhibitory activities of the resulting compounds increased 4.5-55 folds compared to those compounds with the same groups at the R1 (para-) position. In addition, the preferred substituents at the R3 position were Cl or Br, thus compound HS36 exhibited the most potent inhibitory activity (IC50 = 0.15 µM). The molecular docking and site-directed mutation suggest that C128 and N125 are essential for the binding of HS36 and FBPase, which is consistent with the C128-N125-S123 allosteric inhibition mechanism. The reaction enthalpy calculations show that the order of the reactions of compounds with thiol groups at the R3 position is Cl > H > CH3. CoMSIA analysis is consistent with our proposed binding mode. The effect of compounds HS12 and HS36 on glucose production in primary mouse hepatocytes were further evaluated, showing that the inhibition was 71% and 41% at 100 µM, respectively.

CELF6 modulates triple-negative breast cancer progression by regulating the stability of FBP1 mRNA.

BACKGROUND: Triple-negative breast cancer (TNBC) remains a great challenge in clinical treatment due to a shortage of effective therapeutic targets and acquired chemoresistance. Here, we identified the role of an RNA-binding protein, CUG-BP Elav-like family member 6 (CELF6), in the TNBC development and paclitaxel (PTX) chemoresistance. METHODS: Stable CELF6-overexpressing cell lines were established in BT549 and MDA-MB-231 cells. Cell proliferation was determined using cell counting, two-dimensional colony formation, and MTT assay. Meanwhile, cell migration and cell invasion were detected by Transwell assay. Furthermore, the downstream target gene of CELF6 was identified and the direct interaction was further determined by luciferase reporter assay, immunoprecipitation, and RNA pull-down. Additionally, the PTX resistant cell line was established to determine the role of CELF6 in PTX resistance. RESULTS: CELF6 overexpression suppressed cell proliferation, cell migration, and cell invasion. Mechanistically, Fructose-Bisphosphatase 1 (FBP1) was identified as the target gene of CELF6 and stabilized by CELF6 via binding 3'UTR. CELF6 overexpression mediated inhibition in TNBC development was dependent on FBP1. Moreover, CELF6 overexpression increased the sensitivity to PTX treatment. CONCLUSION: CELF6 functions as a tumor suppressor by upregulating FBP 1 expression via stabilizing its mRNA, and thereby inhibits TNBC progression.

Development of disulfide-derived fructose-1,6-bisphosphatase (FBPase) covalent inhibitors for the treatment of type 2 diabetes.

Fructose-1,6-bisphosphatase (FBPase), as a key rate-limiting enzyme in the gluconeogenesis (GNG) pathway, represents a practical therapeutic strategy for type 2 diabetes (T2D). Our previous work first identified cysteine residue 128 (C128) was an important allosteric site in the structure of FBPase, while pharmacologically targeting C128 attenuated the catalytic ability of FBPase. Herein, ten approved cysteine covalent drugs were selected for exploring FBPase inhibitory activities, and the alcohol deterrent disulfiram displayed superior inhibitory efficacy among those drugs. Based on the structure of lead compound disulfiram, 58 disulfide-derived compounds were designed and synthesized for investigating FBPase inhibitory activities. Optimal compound 3a exhibited significant FBPase inhibition and glucose-lowering efficacy in vitro and in vivo. Furthermore, 3a covalently modified the C128 site, and then regulated the N125-S124-S123 allosteric pathway of FBPase in mechanism. In summary, 3a has the potential to be a novel FBPase inhibitor for T2D therapy.

Identification of the New Covalent Allosteric Binding Site of Fructose-1,6-bisphosphatase with Disulfiram Derivatives toward Glucose Reduction.

Fructose 1,6-bisphosphatase (FBPase) has attracted substantial interest as a target associated with cancer and type 2 diabetes. Herein, we found that disulfiram and its derivatives can potently inhibit FBPase by covalently binding to a new C128 allosteric site distinct from the original C128 site in APO FBPase. Further identification of the allosteric inhibition mechanism reveals that the covalent binding of a fragment of 214 will result in the movement of C128 and the dissociation of helix H4 (123-128), which in turn allows S123 to more easily form new hydrogen bonds with K71 and D74 in helix H3 (69-72), thereby inhibiting FBPase activity. Notably, both disulfiram and 212 might moderately reduce blood glucose output in vivo. Therefore, our current findings not only identify a new covalent allosteric site of FBPase but also establish a structural foundation and provide a promising way for the design of covalent allosteric drugs for glucose reduction.

Structural Specificity of Flavonoids in the Inhibition of Human Fructose 1,6-Bisphosphatase.

Liver fructose 1,6-bisphosphatase (FBPase) is a recognized regulatory enzyme of the gluconeogenesis pathway, which has emerged as a valid target to control gluconeogenesis-mediated overproduction of glucose. As such, the management of diabetes with FBPase inhibitors represents a potential alternative for the currently used antidiabetic agents. In this study, the FBPase inhibition of a panel of 55 structurally related flavonoids was tested, through a microanalysis screening system. Then, a subset of seven active inhibitors and their close chemical relatives were further evaluated by molecular dynamics (MD) simulations using a linear interaction energy (LIE) approach. The results obtained showed that D14 (herbacetin) was the most potent inhibitor, suggesting that the presence of -OH groups at the C-3, C-4', C-5, C-7, and C-8 positions, as well as the double bond between C-2 and C-3 and the 4-oxo function at the pyrone ring, are favorable for the intended effect. Furthermore, D14 (herbacetin) is stabilized by a strong interaction with the Glu30 side chain and the Thr24 backbone of FBPase. This is the first investigation studying the in vitro inhibitory effect of a panel of flavonoids against human liver FBPase, thus representing a potentially important step for the search and design of novel inhibitors of this enzyme.

Use of Graph Based Descriptors for Determination of Structural Features Causing Modulation of Fructose-1,6-Bisphosphatase.

Fructose-1,6-bisphosphatase performs a significant function in regulating the blood glucose level in type 2 diabetes by controlling the process gluconeogenesis. In this research work optimal descriptor (graph) based quantitative structural activity relationship studies of a set of 203 fructose-1,6-bisphosphatase has been performed with the help of Monte Carlo optimization. Distribution of compounds into different sets such as training set, invisible training set, calibration set and validation sets resulted in formation of splits. Statistical parameters obtained from quantitative structural activity relationship modeling were good for various designed splits. The statistical parameters such as R2 and Q2 for calibration and validation sets of best split developed were found to be 0.8338, 0.7908 & 0.7920 and 0.7036 respectively. Based on the results obtained for correlation weights, different structural attributes were described as promoters and demoters of the endpoint. Further these structural attributes were used in designing of new fructose-1,6-bisphosphatase inhibitors and molecular docking study was accomplished for the determination of interactions of designed molecules with the enzyme.

Fluoroquinolones suppress gluconeogenesis by inhibiting fructose 1,6-bisphosphatase in primary monkey hepatocytes.

Dysglycemia is one of the most serious adverse events associated with the clinical use of certain fluoroquinolones. The purpose of this study was to investigate the effects of the representative fluoroquinolones moxifloxacin and gatifloxacin on hepatic gluconeogenesis using primary monkey hepatocytes. Glucose production was induced after the cells were incubated for 4 h with 10 mM sodium lactate and 1 mM sodium pyruvate as gluconeogenic substrates. Under these conditions, moxifloxacin and gatifloxacin dose-dependently suppressed gluconeogenesis at concentrations of 100 µM or higher. Transcriptome analysis of rate-limiting enzymes involved in hepatic gluconeogenesis revealed that moxifloxacin and gatifloxacin at a concentration of 1000 µM did not affect the expression of key gluconeogenic enzymes such as phosphoenolpyruvate carboxykinase, glucose 6-phosphatase, and fructose 1,6-bisphosphatase. Furthermore, metabolome analysis, in vitro glucose production assay using additional gluconeogenic substrates, and fructose 1,6-bisphosphatase assay using the cell extracts showed that fluoroquinolones enzymatically suppressed hepatic gluconeogenesis by inhibiting fructose 1,6-bisphosphatase. These inhibitory effects may involve in the clinically relevant dysglycemia associated with fluoroquinolones in human.

GSK343 induces programmed cell death through the inhibition of EZH2 and FBP1 in osteosarcoma cells.

Enhancer of zeste homolog 2 (EZH2) is an important member of the epigenetic regulatory factor polycomb group proteins (PcG) and is abnormally expressed in a wide variety of tumors, including osteosarcoma. Scientists consider EZH2 as an attractive target for the treatment of osteosarcoma and have found many potential EZH inhibitors, such as GlaxoSmithKline 343 (GSK343). It has been reported that GSK343 can be used as an inhibitor in different types of cancer. This study demonstrated that GSK343 not only induced apoptosis by increasing cleaved Casp-3 and poly ADP-ribose polymerase (PARP) expression, but also induced autophagic cell death by inhibiting p62 expression. Apoptosis and autophagic cell death induced by GSK343 were confirmed by the high expression of cleaved caspase-3, LC3-II and transmission electron microscopy. GSK343 inhibited the expression of EZH2 and c-Myc. Additionally, GSK343 inhibited the expression of FUSE binding protein 1 (FBP1), which was identified by its regulatory effects on c-Myc expression. Since c-Myc is a common target of EZH2 and FBP1, and GSK343 inhibited the expression of these proliferation-promoting proteins, a mutual regulatory mechanism between EZH2 and FBP1 was proposed. The knockdown of EZH2 suppressed the expression of FBP1; similarly, the knockdown of FBP1 suppressed the expression of EZH2. These results suggest the mutual regulatory association between EZH2 and FBP1. The knockdown of either EZH2 or FBP1 accelerated the sensitivity of osteosarcoma cells to GSK343. Based on these results, this study clarified that GSK343, an EZH2 inhibitor, may have potential for use in the treatment of osteosarcoma. The underlying mechanisms of the effects of GSK343 are partly mediated by its inhibitory activity against c-Myc and its regulators (EZH2 and FBP1).

Discovery of novel allosteric site and covalent inhibitors of FBPase with potent hypoglycemic effects.

Fructose-1,6-bisphosphatase (FBPase) is an essential enzyme of GNG pathway. Significant advances demonstrate the FBPase plays a critical role in treatment of diabetes. Numerous FBPase inhibitors were developed by targeting AMP site, nevertheless, none of these inhibitors has exhibited suitable potency and druggability. Herein, a new allosteric site (C128) on FBPase was discovered, and several nitrostyrene compounds exhibiting potent FBPase inhibitions were found covalently bind to C128 site on FBPase. Mutagenesis suggest that C128 is the only cysteine that can influence FBPase inhibition, the N125-S124-S123 pathway was most likely involved in allosteric signaling transmission between C128 and active site. However, these nitrostyrenes may bind with multiple cysteine besides C128 in FBPase. To improve pocket selectivity, a series of novel compounds (14a-14n) were re-designed rationally by integrating fragment-based covalent virtual screening and machine-learning-based synthetic complexity evaluation. As expected, the mass spectrometry validated that the proportion of title compounds binding to the C128 in FBPase was significantly higher than that of nitrostyrenes. Notably, under physiological and pathological conditions, the treatment of compounds 14b, 14c, 14i or 14n led to potent inhibition of glucose production, as well as decreased triglyceride and total cholesterol levels in mouse primary hepatocytes. We highlight a novel paradigm that molecular targeting C128 site on FBPase can have potent hypoglycemic effect.

Development of prediction model for fructose- 1,6- bisphosphatase inhibitors using the Monte Carlo method.

Fructose-1,6-bisphosphatase (FBPase) is an enzyme important for regulation of gluconeogenesis, which is a major process in the liver responsible for glucose production. Inhibition of FBPase enzyme causing blockage of the gluconeogenesis process represents a newer scheme in the progress of anti-diabetic drugs. The current research describes the development of hybrid optimal descriptors-based quantitative structure-activity relationship (QSAR) models intended for a set of 62 FBPase inhibitors with the Monte Carlo method. The molecular structures were expressed by the simplified molecular input line entry system (SMILES) notation. Three splits were prepared by random division of the molecules into training set, calibration set and validation set. Statistical parameters obtained from QSAR modelling were good for various designed splits. The best QSAR model showed the following parameters: the values of r2 for calibration set and validation set of the best model were 0.6837 and 0.8623 and of Q2 were 0.6114 and 0.8036, respectively. Based on the results obtained for correlation weights, different structural attributes were described as promoter of the endpoint. Further, these structural attributes were used in designing of new FBPase inhibitors and a molecular docking study was completed for the determination of interactions of the designed molecules with the enzyme.

Fructose-1,6-bisphosphatase Inhibitors: A Review of Recent (2000- 2017) Advances and Structure-Activity Relationship Studies.

Diabetes mellitus, commonly referred to as diabetes, is the 8th leading cause of death worldwide. As of 2015, approximately 415 million people were estimated to be diabetic worldwide, type 2 diabetes being the most common accounting for approximately 90-95% of all diagnosed cases with increasing prevalence. Fructose-1,6-bisphosphatase is one of the important therapeutic targets recently discovered to treat this chronic disease. In this focused review, we have highlighted recent advances and structure-activity relationship studies in the discovery and development of different fructose-1,6-bisphosphatase inhibitors reported since the year 2000.

The FOXC1/FBP1 signaling axis promotes colorectal cancer proliferation by enhancing the Warburg effect.

Aberrant expression of Forkhead box (FOX) transcription factors plays vital roles in carcinogenesis. However, the function of the FOX family member FOXC1 in maintenance of colorectal cancer (CRC) malignancy is unknown. Herein, FOXC1 expression in CRC specimens in The Cancer Genome Atlas (TCGA) cohort was analyzed and validated using immunohistochemistry with a tissue microarray. The effect of FOXC1 expression on proliferation of and glycolysis in CRC cells was assessed by altering its expression in vitro and in vivo. Mechanistic investigation was carried out using cell and molecular biological approaches. Our results showed that FOXC1 expression was higher in CRC specimens than in adjacent benign tissue specimens. Univariate survival analyses of the patients from whom the study specimens were obtained, and validated cohorts indicated that ectopic FOXC1 expression was significantly correlated with shortened survival. Silencing FOXC1 expression in CRC cells inhibited their proliferation and colony formation and decreased their glucose consumption and lactate production. In contrast, FOXC1 overexpression had the opposite effect. Furthermore, increased expression of FOXC1 downregulated that of a key glycolytic enzyme, fructose-1,6-bisphosphatase 1 (FBP1). Mechanistically, FOXC1 bound directly to the promoter regions of the FBP1 gene and negatively regulated its transcriptional activity. Collectively, aberrant FBP1 expression contributed to CRC tumorigenicity, and decreased FBP1 expression coupled with increased FOXC1 expression provided better prognostic information than did FOXC1 expression alone. Therefore, the FOXC1/FBP1 axis induces CRC cell proliferation, reprograms metabolism in CRCs, and constitutes potential prognostic predictors and therapeutic targets for CRC.

In silico screening of a novel scaffold for fructose-1,6-bisphosatase (FBPase) inhibitors.

Fructose-1, 6-bisphosphatase (FBPase) has been regarded as an attractive drug target to control blood glucose against Type 2 diabetes (T2D). In this study, by using the strategy of pharmacophore-based virtual screening, a novel scaffold inhibitor targeted the AMP allosteric site of human liver FBPase were screened, their inhibitory activities were further tested. The experimental results showed that compound H27 exhibited high inhibitory activities with the IC50 value of 5.3 µM. Therefore, compound H27 was chosen as the probe molecule, it's possible binding conformation targeted into FBPase was identified by using DOX2.0 strategy. The importance of key residues (T27, T31, K112 and R140) in allosteric site of FBPase for the binding inhibitors were validated by mutation experiments. The agreement between theory and experiment suggest that the interactional information of FBPase and inhibitors (H27) were reliable. On basis of these rational interactional information, the compound H29 was further designed to exhibit more potential FBPase inhibition (IC50 = 2.5 µM).

Current anti-diabetic agents and their molecular targets: A review.

Diabetes mellitus is a medical condition characterized by the body's loss of control over blood sugar. The frequency of diagnosed cases and consequential increases in medical costs makes it a rapidly growing chronic disease that threatens human health worldwide. In addition, its unnerving statistical projections are perilous to both the economy of the nation and man's life expectancy. Type-I and type-II diabetes are the two clinical forms of diabetes mellitus. Type-II diabetes mellitus (T2DM) is illustrated by the abnormality of glucose homeostasis in the body, resulting in hyperglycemia. Although significant research attention has been devoted to the development of diabetes regimens, which demonstrates success in lowering blood glucose levels, their efficacies are unsustainable due to undesirable side effects such as weight gain and hypoglycemia. Over the years, heterocyclic scaffolds have been the basis of anti-diabetic chemotherapies; hence, in this review we consolidate the use of bioactive scaffolds, which have been evaluated for their biological response as inhibitors against their respective anti-diabetic molecular targets over the past five years (2012-2017). Our investigation reveals a diverse target set which includes; protein tyrosine phosphatase 1 B (PTP1B), dipeptidly peptidase-4 (DPP-4), free fatty acid receptors 1 (FFAR1), G protein-coupled receptors (GPCR), peroxisome proliferator activated receptor-γ (PPARγ), sodium glucose co-transporter-2 (SGLT2), α-glucosidase, aldose reductase, glycogen phosphorylase (GP), fructose-1,6-bisphosphatase (FBPase), glucagon receptor (GCGr) and phosphoenolpyruvate carboxykinase (PEPCK). This review offers a medium on which future drug design and development toward diabetes management may be modelled (i.e. optimization via structural derivatization), as many of the drug candidates highlighted show promise as an effective anti-diabetic chemotherapy.

Fructose-1,6-bisphosphatase inhibitors: A new valid approach for management of type 2 diabetes mellitus.

The rising incidence of diabetes and confines allied with clinical therapies emphasized the need to explore new molecular targets to develop novel, effective and safer antihyperglycemic agents. Excessive endogenous glucose production by gluconeogenesis is a primary determinant of hyperglycemia in patients with type 2 diabetes. But not even a single current medication acts directly to reduce gluconeogenesis. Fructose-1,6-bisphosphatase (FBPase), a well recognized rate controlling enzyme of gluconeogenesis, has emerged as legitimate molecular level target to control gluconeogenesis mediated glucose overproduction and its inhibitors are likely to fulfill an unmet medical need. In this compilation various chemical classes of FBPase inhibitors have been reviewed which mainly acts through uncompetitive and non-competitive manner. A detailed account on structure activity relationship studies of inhibitors have been presented along with their molecular level interactions at binding sites of enzyme. Three Dimensional Quantitative Structure Activity relationship (3D-QSAR) studies, performed to optimize the lead, have been summarized at some places. In this assemblage, FBPase inhibitors patented in past have been compiled in tabular form. This review highlights the new insight into the therapeutic utility of FBPase inhibitors and their potential as a new class of antidiabetic drugs.

Structures of Leishmania Fructose-1,6-Bisphosphatase Reveal Species-Specific Differences in the Mechanism of Allosteric Inhibition.

The gluconeogenic enzyme fructose-1,6-bisphosphatase has been proposed as a potential drug target against Leishmania parasites that cause up to 20,000-30,000 deaths annually. A comparison of three crystal structures of Leishmania major fructose-1,6-bisphosphatase (LmFBPase) along with enzyme kinetic data show how AMP acts as an allosteric inhibitor and provides insight into its metal-dependent reaction mechanism. The crystal structure of the apoenzyme form of LmFBPase is a homotetramer in which the dimer of dimers adopts a planar conformation with disordered "dynamic loops". The structure of LmFBPase, complexed with manganese and its catalytic product phosphate, shows the dynamic loops locked into the active sites. A third crystal structure of LmFBPase complexed with its allosteric inhibitor AMP shows an inactive form of the tetramer, in which the dimer pairs are rotated by 18° relative to each other. The three structures suggest an allosteric mechanism in which AMP binding triggers a rearrangement of hydrogen bonds across the large and small interfaces. Retraction of the "effector loop" required for AMP binding releases the side chain of His23 from the dimer-dimer interface. This is coupled with a flip of the side chain of Arg48 which ties down the key catalytic dynamic loop in a disengaged conformation and also locks the tetramer in an inactive rotated T-state. The structure of the effector site of LmFBPase shows different structural features compared with human FBPases, thereby offering a potential and species-specific drug target.