High-resolution crystal structure of LpqH, an immunomodulatory surface lipoprotein of Mycobacterium tuberculosis reveals a distinct fold and a conserved cleft on its surface.

Tuberculosis, caused by Mycobacterium tuberculosis, is predominantly a disease of the lungs acquired by inhaling mycobacteria from infected individuals via airborne droplets. In order to facilitate their entry into the alveolar macrophages, mycobacteria have a collection of pathogen-associated molecular patterns (PAMPs) on their surface that are known to detect certain pattern recognition receptors present on the surface of host cells. A major group of these PAMPs includes mycobacterial lipoproteins, of which, the 19 kDa surface antigen LpqH, has been reported to play a critical role in both host-pathogen interactions as well as pleiotropic immune regulation. Despite its crucial involvement in tuberculosis, the detailed structure-function relationship of this protein remains to be explored. Here, we report the high-resolution crystal structure of the non-acylated LpqH (LpqH48-159) at a resolution of 1.26 Å, which adopts a unique fold. Flow cytometry-based experiments show that the protein can bind and induce apoptosis in PMA-activated human monocytic cell line THP-1, indicative of the preservation of functionality of the protein. Furthermore, analysis of conservation of LpqH sequences from Mycobacterium species reveals a patch of conserved residues on the surface which may play a role in its binding partner recognition and hence in host-pathogen interaction.

Characterization of an alcohol acetyltransferase GcAAT responsible for the production of antifungal volatile esters in endophytic Geotrichum candidum PF005.

Alcohol acetyltransferases (AATs) are a group of enzymes that catalyze the formation of esters from different alcohols and acetyl-CoA. However, these enzymes are not well characterized with regard to synthesis of antifungal compounds. The present study aims to investigate the AAT enzyme from Geotrichum candidum PF005, an endophytic yeast-like fungus that emits fruity scented antifungal volatiles, primarily comprising of acetate esters. After PCR-based cloning of the GcAAT gene, the encoded enzyme was characterized structurally through in silico methods and functionally via heterologous expression in Saccharomyces cerevisiae. In native host, the single copy GcAAT gene exhibited induced expression upon supplementation with metabolic precursors, like L-leucine (Leu) or α-ketoisocaproate (α-KIC). Docking studies using the modelled structure of GcAAT revealed differential but favourable binding interactions for three alcohol substrates (i.e., isoamyl alcohol, isobutyl alcohol and 2-phenylethanol) and the co-substrate acetyl-CoA. Binding sites for both substrate and co-substrate are found to be located inside a tunnel identified in the structure, wherein the H208 of the acetyltransferase conserved motif HXXXD was found at a hydrogen bond distance from the substrate. Functional complementation of GcAAT in S. cerevisiae AAT knockout strain caused 32% decrease in dry biomass weight of the test phytopathogenic fungus, Rhizoctonia solani as compared to the control (AAT knockout strain with empty plasmid) after 72 h of incubation due to the emitted volatiles. When the transformed yeast cells were fed with Leu and α-KIC, the relative abundance of the isoamyl acetate ester increased by 21% and 48%, respectively as compared to the control (without precursor). Further analysis documented that volatiles from α-KIC fed GcAAT transformant exhibited 58% higher antifungal activity against the test fungus R. solani than the control, engendered by increased oxidative stress that led to distorted mycelial morphology and increased hyphal branching. Together, the augmented antifungal effect displayed by the GcAAT expressing S. cerevisiae AAT knockout strain is clearly attributable to the acetate esters, especially isoamyl acetate, which are inherently produced in endophytic G. candidum PF005 as antifungal volatiles.

The C-terminal end of mycobacterial HadBC regulates AcpM interaction during the FAS-II pathway: a structural perspective.

Comprehending the molecular strategies employed by Mycobacterium tuberculosis (Mtb) in FAS-II regulation is of paramount significance for curbing tuberculosis progression. Mtb employs two sets of dehydratases, namely HadAB and HadBC (ß-hydroxyacyl acyl carrier protein dehydratase), for the regulation of the fatty acid synthase (FAS-II) pathway. We utilized a sequence similarity network to discern the basis for the presence of two copies of the dehydratase gene in Mtb. This analysis groups HadC and HadA in different clusters, which could be attributed to the variability in their physiological role with respect to the acyl chain uptake. Our study reveals structural details pertaining to the crystal structure of the last remaining enzyme of the FAS-II pathway. It also provides insights into the highly flexible hot-dog helix and substrate regulatory loop. Additionally, mutational studies assisted in establishing the role of the C-terminal end in HadC of HadBC in the regulation of acyl carrier protein from Mtb-mediated interactions. Complemented with surface plasmon resonance and molecular dynamics simulation studies, the present study provides the first evidence of the molecular mechanisms involved in the differential binding affinity of the acyl carrier protein from Mtb towards both mtbHadAB and mtbHadBC.

Fluorescently labelled thioacetazone for detecting the interaction with Mycobacterium dehydratases HadAB and HadBC.

Thioacetazone (TAC) used to be a highly affordable, bacteriostatic anti-TB drug but its use has now been restricted, owing to severe side-effects and the frequent appearance of the TAC resistant M. tuberculosis strains. In order to develop new TAC analogues with fewer side-effects, its target enzymes need to be firmly established. It is now hypothesized that TAC, after being activated by a monooxygenase EthA, binds to the dehydratase complex HadAB that finally leads to a covalent modification of HadA, the main partner involved in dehydration. Another dehydratase enzyme, namely HadC in the HadBC complex, is also thought to be a possible target for TAC, for which definitive evidence is lacking. Herein, using a recently exploited azido naphthalimide template attached to thioacetazone and adopting a photo-affinity based labelling technique, coupled with electrophoresis and in-gel visualization, we have successfully demonstrated the involvement of these enzymes including HadBC along with a possible participation of an alternate mycobacterial monooxygenase MymA. In silico studies also revealed strong interactions between the TAC-probe and the concerned enzymes.

Residues at the interface between zinc binding and winged helix domains of human RECQ1 play a significant role in DNA strand annealing activity.

RECQ1 is the shortest among the five human RecQ helicases comprising of two RecA like domains, a zinc-binding domain and a RecQ C-terminal domain containing the winged-helix (WH). Mutations or deletions on the tip of a ß-hairpin located in the WH domain are known to abolish the unwinding activity. Interestingly, the same mutations on the ß-hairpin of annealing incompetent RECQ1 mutant (RECQ1T1) have been reported to restore its annealing activity. In an attempt to unravel the strand annealing mechanism, we have crystallized a fragment of RECQ1 encompassing D2-Zn-WH domains harbouring mutations on the ß-hairpin. From our crystal structure data and interface analysis, we have demonstrated that an α-helix located in zinc-binding domain potentially interacts with residues of WH domain, which plays a significant role in strand annealing activity. We have shown that deletion of the α-helix or mutation of specific residues on it restores strand annealing activity of annealing deficient constructs of RECQ1. Our results also demonstrate that mutations on the α-helix induce conformational changes and affects DNA stimulated ATP hydrolysis and unwinding activity of RECQ1. Our study, for the first time, provides insight into the conformational requirements of the WH domain for efficient strand annealing by human RECQ1.

Mycobacterial crypto-AcpM as a tool to investigate the consequence of drug binding on its key FAS II partner enzyme HadAB.

Background Mycobacterial FASII pathway is governed by the Protein-Protein Interaction mediated dynamics existent between Acyl Carrier Protein and its partner enzymes. The dehydratase HadAB, involved in the third step of FASII synthesis has remained a key target of drugs like Thiacetazone (TAC) and its consequence on AcpM binding is yet to be deciphered. Owing to the transient nature of these interactions, analysing their implications as a drug target has been exhausting. Methods In this context, we have developed an in vitro method to study the effect of thiocarbamide-containing compounds, TAC and SPA0355 (a thiourea analogue) against mycobacterial HadAB. Additionally, by utilizing crypto-ACP (NBD-tagged Acyl Carrier Protein) as a tool of our choice, we attempted at exploring the effect of TAC and SPA0355 on mycobacterial HadAB. Results SPA0355 behaves at par with TAC and undergoes activation in the presence of monooxygenase EthA thus, bringing about a covalent modification in HadA subunit of HadAB. The crypto-ACP method provides insights into the altered substrate housing capability in HadAB associated with the impediment of its AcpM mediated functionality; an outcome attributed to the repercussions associated with the binding of the aforementioned thiourea compounds. Conclusion This investigation has assisted in unveiling a two-step mechanism undertaken by AcpM for interacting with its corresponding partner protein during acyl chain transfer. General significance This study highlights the alterations brought about by drug binding in the interplay between ACP and HadAB. Additionally, this work for the first time establishes the role of SPA0355 as a promising drug candidate against dehydratase HadAB.

Silver Nanodot Decorated Dendritic Copper Foam As a Hydrophobic and Mechano-Chemo Bactericidal Surface.

The present work investigates the time-dependent antibacterial activity of the silver nanodot decorated dendritic copper foam nanostructures against Escherichia coli (Gram-negative) and Bacillus subtilis (Gram-positive) bacteria. An advanced antibacterial and antifouling surface is fabricated utilizing the collective antibacterial properties of silver nanodots, chitosan, and dendritic copper foam nanostructures. The porous network of the Ag nanodot decorated Cu foam is made up of nanodendrites, which reduce the wettability of the surface. Hence, the surface exhibits hydrophobic nature and inhibits the growth of bacterial flora along with the elimination of dead bacterial cells. The fabricated surface exhibits a water contact angle (WCA) of 158.7 ± 0.17°. Specifically, we tested the fabricated material against both the Gram-positive and Gram-negative bacterial models. The antibacterial activity of the fabricated surface is evident from the growth inhibition percentage of bacterial strains of Escherichia coli (72.30 ± 0.60%) and Bacillus subtilis (48.30 ± 1.71%). The micrographs obtained from scanning electron microscopy (SEM), transmission electron microscopy (TEM), and atomic force microscopy (AFM) of the treated cells show the damaged cellular structures of the bacteria, which is strong evidence of successful antibacterial action. The antibacterial effect can be attributed to the synergistic mechano-chemo mode of action involving mechanical disruption of the bacterial cell wall by the nanoprotrusions present on the Cu dendrites along with the chemical interaction of the Ag nanodots with vital intracellular components.

Design and development of a machine for continuous popping and puffing of grains.

Popping/puffing have been traditionally practiced for enhancing storage life, improving organoleptic properties and ease of incorporation in ready-to-eat-foods. Currently, batch type sand and electric popping/puffing machines involving conduction mode of heat transfer are employed. The major drawbacks of these methods are high-energy consumption, scorching of grains, non-uniform product quality, contamination (by sand/ash) and problems in scale-up. Since fluidization is known to increase heat and mass transfer, a continuous fluidized popping/puffing machine (capacity 10-20 kg/h) involving convective mode of heat transfer is designed/developed. Hot-flue gas generating from burning of LPG was used as the eco-friendly fuel. Process parameters such as expansion ratio, fluidization velocity, terminal velocity, carry over velocity, bulk density and voidage were estimated for un-popped and popped/puffed rice, maize, jowar (sorghum) and paddy. Fluidization and carry over velocities for these grains were in the range of 4.18-5.78 m/s and 2.15-6.18 m/s, respectively. Based on the terminal velocity of the grains and volumetric air flow rate of the blower, fluidization chamber diameter was arrived. Chamber diameter of 0.15 m was found to be sufficient to generate required air velocity of 6.89 m/s which met the fluidization and carry over velocities of popped/puffed grains. The designed fluidization chamber was analyzed for heat and mass transfer during popping/puffing. Convective heat and mass transfer coefficients were estimated to be in the range of 103-187 W/m2 °C and 0.124-0.162 m/s, respectively. Theoretical values for total heat and mass transfer were similar to the experimental values.

Characterization of two sugar transporters responsible for efficient xylose uptake in an oleaginous yeast Candida tropicalis SY005.

Microbial conversion of lignocellulosic feedstock to the target bioproduct requires efficient assimilation of its constituent sugars, a large part of which comprises of glucose and xylose. This study aims to identify and characterize sugar transporters capable of xylose uptake in an oleaginous strain of the industrially relevant yeast Candida tropicalis. In silico database mining resulted in two sugar transporter proteins- CtStp1 and CtStp2, containing conserved amino acid residues and motifs that have been previously reported to be involved in xylose transport in other organisms. Several softwares predicted the likelihood of 10-12 transmembrane (TM) helices to be present in both the Stps, while molecular modelling showed 12 TM helices that were organized into a typical structure found in the major facilitator superfamily of transporters. Docking with different sugars also predicted favorable interactions. Heterologous expression in a Saccharomyces cerevisiae strain harboring functional xylose metabolic genes validated the broad substrate specificity of the two Stps. Each transporter supported prominent growth of recombinant S. cerevisiae strains on six sugars including xylose at various concentrations. Expression of CtSTP1 and CtSTP2 along with the xylose metabolic genes in yeast transformants grown in presence of xylose was confirmed by transcript detection. Growth curve and sugar consumption profiles revealed uptake of both glucose and xylose simultaneously by the recombinant yeast strains, though CtStp1 showed relatively less effect of glucose repression in mixed sugars and was a better transporter of xylose than CtStp2. Such glucose-xylose utilizing efficient transporters can be effective tools for developing co-fermenting yeasts through genetic engineering in future, with noteworthy applications in renewable biomass utilization.

Identification and functional characterization of a lipid droplet protein CtLDP1 from an oleaginous yeast Candida tropicalis SY005.

Proteins residing in lipid droplets (LDs) of organisms exhibit diverse physiological roles. Since the LD proteins of yeasts are largely unexplored, we have identified a putative LD protein gene, CtLDP1 in the oleaginous yeast Candida tropicalis SY005 and characterized its function. The increased lipid accumulation in SY005 could be correlated with enhanced (~2.67-fold) expression of the CtLDP1 after low-nitrogen stress. The N-terminal transmembrane domain similar to perilipin proteins and the amphipathic α-helices predicted in silico, presumably aid in targeting the CtLDP1 to LD membranes. Heterologous expression of CtLDP1-mCherry fusion in Saccharomyces cerevisiae revealed localization in LDs, yet the expression of CtLDP1 did not show significant effect on LD formation in transformed cells. Molecular docking showed favourable interactions of the protein with sterol class of molecules, but not with triacylglycerol (TAG); and this was further experimentally verified by co-localization of the mCherry-tagged protein in TAG-deficient (but steryl ester containing) LDs. While oleic acid supplementation caused coalescence of LDs into supersized ones (average diameter = 1.19 ± 0.12 µm; n = 160), this effect was suppressed due to CtLDP1 expression, and the cells mostly exhibited numerous smaller LDs (average diameter = 0.46 ± 0.05 µm; n = 160). Moreover, CtLDP1 expression in pet10Δ knockout strain of S. cerevisiae restored multiple LD formation, indicating functional complementation of the protein. Overall, this study documents functional characterization of an LD-stabilizing protein from an oleaginous strain of Candida genus for the first time, and provides insights on the characteristics of LD proteins in oleaginous yeasts for future metabolic engineering.

RUVBL1/RUVBL2 ATPase Activity Drives PAQosome Maturation, DNA Replication and Radioresistance in Lung Cancer.

RUVBL1 and RUVBL2 (collectively RUVBL1/2) are essential AAA+ ATPases that function as co-chaperones and have been implicated in cancer. Here we investigated the molecular and phenotypic role of RUVBL1/2 ATPase activity in non-small cell lung cancer (NSCLC). We find that RUVBL1/2 are overexpressed in NSCLC patient tumors, with high expression associated with poor survival. Utilizing a specific inhibitor of RUVBL1/2 ATPase activity, we show that RUVBL1/2 ATPase activity is necessary for the maturation or dissociation of the PAQosome, a large RUVBL1/2-dependent multiprotein complex. We also show that RUVBL1/2 have roles in DNA replication, as inhibition of its ATPase activity can cause S-phase arrest, which culminates in cancer cell death via replication catastrophe. While in vivo pharmacological inhibition of RUVBL1/2 results in modest antitumor activity, it synergizes with radiation in NSCLC, but not normal cells, an attractive property for future preclinical development.

Electrodeposited Cu2O Nanopetal Architecture as a Superhydrophobic and Antibacterial Surface.

Bacterial infections being sporadic and uncontrollable demands an urgent paradigm shift in the development of novel antibacterial agents. This work involves the fabrication of Cu2O nanopetals over copper foil that show superlative antibacterial and superhydrophobic properties. A superhydrophobic surface has been fabricated using the electrochemical deposition (ECD) method. Here, it is aimed to establish the superior antibacterial activity as an outcome of the inherent superhydrophobic property of the as-fabricated nanostructures. The present study finds that the elevated value of the water contact angle (154 ± 0.6°) does not allow proper bacterial adhesion, and it is immune from the possibility of biofouling. Specifically, two kinds of bacterial strains have been tested and the time response of the antibacterial activity has been studied over a period of 12 h, taking DH5α Escherichia coli as a Gram-negative model and Bacillus subtilis 168 as a Gram-positive model. Higher antibacterial effects were observed for the Gram-negative model (E. coli) owing to its simplistic cell wall structure which facilitates the easy diffusion of Cu+ ions into the bacterial membrane. The simplicity of the developed method of fabrication along with the superlative superhydrophobic nature and excellent antibacterial property of the material, owing to its synergistic biophysical and biochemical modes of biocidal action, establishes its viability in many applications.

Naphthalimide-Based Template for Inhibitor Screening via Cross-Linking and In-Gel Fluorescence: A Case Study against HCA II.

We describe a rapid electrophoresis-based method for profiling of carbonic anhydrase inhibitors. In addition to the pharmacophore moiety intended for reversible interaction with a target enzyme, a fluorescent template with a built-in azide group for photoaffinity labeling is also included as a part of the inhibitor design. Following incubation and irradiation, gel electrophoresis with visualization under UV allows assessment of the efficiency of cross-linking. The relative efficiency of cross-linking of various probes can be regarded as a reflection of their inhibition potencies, an assumption supported by the trend in their IC50/K i values. The method has the advantage of being applicable to impure enzyme preparations and also can be used to screen several inhibitors including their promiscuity in parallel in a short time as has been currently demonstrated with HCA II.

Probing into Therapeutic Anti-cancer Potential of Apigenin: Recent Trends and Future Directions.

BACKGROUND: Natural products represent a therapeutic option for the treatment of inflammation- associated diseases. Flavonoids which are one of the special categories of such natural products, have previously shown promising therapeutic potential. OBJECTIVES: The current review discusses the synthetic preview and anti-inflammatory potential of apigenin along with the underlying molecular mechanism in chronic human diseases especially cancer. In addition, the relevant patents on the therapeutic potential of apigenin have also been mentioned. METHODS: A literature search was carried out using PubMed/Science, Google Scholar, etc. which was further expended by the different combination of keywords: apigenin, inflammation, mechanism, therapeutic potential, cancer, etc. Patent information was retrieved by searching the key terms: apigenin, inflammation, therapeutic potential from various databanks including Espacenet, Google Patents, Free Patents Online and Mendeley of WIPO, USPTO, SIPO, JPO, KIPO and EPO databases. RESULTS: A total of 76 references have been found relevant with the theme of the manuscript. These citations have described recent ongoing advances in the area of inflammation and cancer with respect to apigenin. CONCLUSION: Studies related to the anti-inflammatory and anticancer potential of apigenin have been explored through this review article. Moreover, the patent analysis of apigenin has further strengthened its therapeutic role. Probing into the therapeutic properties of apigenin, further adds value to this molecule in terms of its downregulation of major inflammatory and cancer-associated signaling pathways. The article would simultaneously assist the scientific community to precisely understand the role of apigenin and design novel anti-cancer therapies.

Jumonji Inhibitors Overcome Radioresistance in Cancer through Changes in H3K4 Methylation at Double-Strand Breaks.

We have uncovered a role for Jumonji inhibitors in overcoming radioresistance through KDM5B inhibition. Pharmacological blockade of Jumonji demethylases with JIB-04 leads to specific accumulation of H3K4me3 at sites marked by γH2AX and impaired recruitment of DNA repair factors, preventing resolution of damage and resulting in robust sensitization to radiation therapy. In DNA-repair-proficient cancer cells, knockdown of the H3K4me3 demethylase KDM5B, but not other Jumonji enzymes, mimics pharmacological inhibition, and KDM5B overexpression rescues this phenotype and increases radioresistance. The H3K4me3 demethylase inhibitor PBIT also sensitizes cancer cells to radiation, while an H3K27me3 demethylase inhibitor does not. In vivo co-administration of radiation with JIB-04 significantly prolongs the survival of mice with tumors even long after cessation of treatment. In human patients, lung squamous cell carcinomas highly expressing KDM5B respond poorly to radiation. Thus, we propose the use of Jumonji KDM inhibitors as potent radiosensitizers.

Bioactives-retained non-glutinous noodles from nixtamalized Dent and Flint maize.

Nixtamalization is a well-known pre-treatment technique in the tortilla industry. Nixtamalized maize (nixtamal) is known for its modified physicochemical as well as nutritional attributes. In the present study, two types of nixtamalization processes (traditional and ecological) were employed for the development of whole-grain-maize-based noodles using Dent and Flint maize genotypes. Results showed that ecological nixtamalization had resulted in better cooking and textural qualities of noodles compared to the one prepared traditionally. Dent maize noodles from traditional and ecological nixtamalization had lower retention of phenolics (40 and 64%, respectively) whereas, Flint maize noodles retained 50 and 66% phenolics, respectively. Dent maize noodles had undergone phenolics loss of 5-6% on cooking while those of Flint maize lost only 2%. Ecological nixtamalization maintained the pH of the cooking liquor within an acidic-neutral range and yielded noodle with higher retention of phenolics whereas, the traditional process negatively affected the antioxidant compounds and their properties.

Antioxidative free and bound phenolic constituents in botanical fractions of Indian specialty maize (Zea mays L.) genotypes.

Specialty maize genotypes viz. QPM (quality protein maize), Baby corn, Popcorn and Sweet corn, which are usually consumed in whole forms can be good supplements of phenolic antioxidants. Botanical fractions of these maize genotypes were analyzed to explore the distribution of free and bound phenolics. HPLC and ESI-MS/MS results indicated the presence of vanillic, syringic, p-hydroxybenzoic, caffeic, p-coumaric, ferulic and isoferulic acids along with cyanidin-3-O-glucoside, kaempferol and quercetin. Germs of maize samples contained significantly higher free phenolics than pericarps, whereas, pericarps contained 74-83% of bound ones. QPM and Popcorn contained only 3% free phenolics whereas, Baby corn and Sweet corn had 14-17%. Unlike in peroxide scavenging and reducing capacity, anti-radical capacity of free phenolics of germs was significantly higher than that of pericarps. Free phenolics contributed 0.2-1.65%, 2-5% and 42-49% in anti-radical, peroxide scavenging and reducing capacity, respectively. Among lipophilic tocochromanols γ-tocopherol was the most abundant isomer in the samples among which Sweet corn contained the most (84.2 µg/g). Data showed that specialty maize genotypes are rich sources of hydrophilic and lipophilic bioactives and are natural antioxidants.

Design, synthesis and characterization of dual inhibitors against new targets FabG4 and HtdX of Mycobacterium tuberculosis.

Herein, we present dual inhibitors of new targets FabG4 and HtdX for the first time. In this work, eight compounds have been designed, synthesized, characterized and evaluated for bio-activities. Amongst them, six compounds have shown inhibitory activities. Three of them (12-14) demonstrate dual inhibition of both FabG4 and HtdX at low micromolar concentration. In addition, the dual inhibitors show good anti-mycobacterial properties against both planktonic growth and biofilm culture of Mycobacterium species. This study is an important addition to tuberculosis drug discovery because it explores two new enzymes as drug targets and presents their dual inhibitors as good candidates for pre-clinical trials.

Inhibition of M. tuberculosis ß-ketoacyl CoA reductase FabG4 (Rv0242c) by triazole linked polyphenol-aminobenzene hybrids: comparison with the corresponding gallate counterparts.

Herein we report six novel triazole linked polyphenol-aminobenzene hybrids (3-8) as inhibitors of Mycobacterium tuberculosis FabG4 (Rv0242c), a less explored ß-ketoacyl CoA reductase that has immense potential to be the future anti-tuberculosis drug target due to its possible involvement in drug resistance and latent infection. Novel triazole linked polyphenol-aminobenzene hybrids have been synthesized, characterized and evaluated for their inhibitory activity against FabG4. All of them inhibit FabG4 at low micromolar concentrations. In silico docking study has been carried out to explain the experimental findings. A comparative study of these new inhibitors with previously reported gallate counterparts leads to structure-activity relations (SAR) of substituent linked to N-1 of triazole ring.

Isolation and characterization of extracellular polysaccharide Thelebolan produced by a newly isolated psychrophilic Antarctic fungus Thelebolus.

The present investigation is on a newly isolated psychrophilic Antarctic filamentous Ascomycetous fungus that has been identified as Thelebolus sp. and given the designation of Thelebolus sp. IITKGP-BT12. The culture was primarily identified through morphological studies, and was further confirmed by 18S rRNA sequencing (GenBank Accession No. KC191572), which revealed its close relatedness with Thelebolus microsporus. The exopolysaccharide (EPS) produced (1.94 g L(-1)) by the fungus was isolated, purified and characterized as glucan having an average molecular mass of 5×10(5)Da. The structure of EPS was determined by gas chromatography with tandem mass spectrometry (GC-MS/MS), Fourier transform infrared spectroscopy (FT-IR) and nuclear magnetic resonance (NMR) studies ((1)H, (13)C and HSQC). NMR analysis indicated the presence of (1→3)-linked ß-d-glucan backbone with (1→6)-linked branches of ß-d-glucopyranosyl units. Antiproliferative activity of EPS was demonstrated in B16-F0 cells, with IC50 of 275.42 µg m L(-1). Flow cytometry analysis and DNA fragmentation studies revealed that the cytotoxic action of the EPS mediated apoptosis in cancer cells. This is the first ever report on bioactive EPS thelebolan from Thelebolus sp.