Comparison of Creatine Kinase-MB, Troponin-I, Troponin-T, Triiodothyronine, Thyroxine, Apoprotein-B and Homocysteine between Non-smokers, Smokers at High Altitudes, and Smokers at Sea Level.

Earlier diagnosis of heart disease can occur via awareness of biochemical changes. Keeping this in view, we wanted to determine if there was any difference between biochemical heart parameters between non-smokers (the control group), smokers who live at a high altitude, or smokers who live at sea level. There were 180 participants categorised into three groups, A, B, and C, depending upon their smoking/non-smoking classification, or distance from sea level. Blood samples were taken as per requirements to check levels of creatine kinase-MB, troponin-I, troponin-T, Triiodothyronine (T3), Thyroxine (T4), Apolipoprotein B (apo-B), and homocysteine, and subjected to enzyme-linked immunoassay (ELISA) investigations. Creatine kinase-MB, troponin-I, troponin-T, T3, thyroxine, apoprotein-B, and homocysteine all exhibited a noteworthy difference (p≤0.01) when compared between non-smokers and smokers (either at a high altitude or sea level), but only troponin I and T3 showed a noteworthy difference when compared between smokers at a high altitude versus at sea level (p≤0.01) as follows: Creatine kinase-MB, p=0.434; troponin-I, troponin-T, p=0.208; T3, p≤0.01; thyroxine, p=0.190; apoprotein-B, p=0.008; and homocysteine, p=0.039. It has been found that significant differences exist between smokers and non-smokers regarding cardiovascular (CV) pathology, whether the person resides at a high altitude or sea level. However, additional studies should be performed to find the correlation between smokers at a high altitude versus and smokers at sea level, which can change the treatment methods at high altitudes and pave the way for finding new medicines.

Klebsiella oxytoca: an efficient pyrene-degrading bacterial strain isolated from petroleum-contaminated soil.

Polycyclic aromatic hydrocarbons (PAHs) are the hazardous xenobiotic agents of oil production. One of the methods to eliminate hazardous compounds is bioremediation, which is the most efficient and cost-effective method to eliminate the harmful byproducts of crude petroleum processing. In this study, five pure bacterial isolates were isolated from petroleum-contaminated soil, four of which showed a robust growth on the PAH pyrene, as a sole carbon source. Various methods viz mass spectroscopy, biochemical assays, and 16S RNA sequencing employed to identify the isolates ascertained the consistent identification of Klebsiella oxytoca by all three methods. Scanning electron microscopy and Gram staining further demonstrated the characterization of the K. oxytoca. High-performance liquid chromatography of the culture supernatant of K. oxytoca grown in pyrene containing media showed that the cells started utilizing pyrene from the 6th day onwards and by the 12th day of growth, 70% of the pyrene was completely degraded. A genome search for the genes predicted to be involved in pyrene degradation using Kyoto Encyclopedia of Genes and Genomes (KEGG) confirmed their presence in the genome of K. oxytoca. These results suggest that K. oxytoca would be a suitable candidate for removing soil aromatic hydrocarbons.

Antimycobacterial and Anticancer Properties of Myrtus communis Leaf Extract.

BACKGROUND: Plant-derived products or extracts are widely used in folk/traditional medicine to treat several infections, ailments, or disorders. A well-known medicinal herb, Myrtus communis is an evergreen fragrant plant native to the Mediterranean region that has been used for ages in traditional medicine around the world. MATERIALS AND METHODS: The microplate alamarBlue assay and the well diffusion method were used to evaluate the zone of inhibition and MIC, respectively. The double-disc diffusion method was used to investigate the synergy between antibiotics and the extract. The crystal violet method was used to investigate biofilm development. The SulphoRhodamine-B assay and DNA flow cytometry were used to investigate the proliferation and subsequent distribution of cells among different phases of the cell cycle. The apoptotic and necrotic phases of the cancer cells were examined using flow cytometry in conjunction with Annexin V-FITC/PI labeling. Using the IBM SPSS statistical program, a one-way ANOVA with Tukey's post hoc test was employed for statistical analysis. RESULTS: The ethanolic leaf extract of M. communis showed a strong growth inhibition effect (zone of inhibition: 20.3 ± 1.1-26.3 ± 2.5 mm, MIC: 4.88-312.5 µg/mL, and MBC: 39.07-1250 µg/mL) against several rapidly growing and slow-growing mycobacterial strains in a dose-dependent manner. Damage to the cell wall of bacterial cells was determined to be the cause of the antimycobacterial action. The extract inhibited biofilm formation (MBIC of 9.7 µg/mL) and eradicated already-formed mature and ultra-mature biofilms of M. smegmatis, with MBEC values of 78 µg/mL and 156 µg/mL, respectively. Additionally, the extract exhibited potent anticancer effects against diverse cancer cell lines of the breast (MCF-7), liver (HepG2), cervix (HeLa), and colon (HCT116) (IC50 for HCT116: 83 ± 2.5, HepG2: 53.3 ± 0.6, MCF-7: 41.5 ± 0.6, and HeLa: 33.3 ± 3.6) by apoptosis after arresting the cells in the G1 phase of the cell cycle. CONCLUSIONS: These results suggest that M. communis leaf extract is a potential source of secondary metabolites that could be further developed as potential anticancer and antimycobacterial agents to treat diverse types of cancers and mycobacterial infections.

Antibacterial and Antibiofilm Activities of ß-Lapachone by Modulating the Catalase Enzyme.

BACKGROUND: Bacterial infections constantly have a large impact on public health, because of increased rates of resistance and reduced frequency of development of novel antibiotics. The utility of conventional antibiotics for treating bacterial infections has become increasingly challenging. The aim of the study was to assess the antibacterial effect of ß-Lapachone (ß-Lap), a novel synthetic compound. METHODS: The antibacterial activity of the ß-Lap compound was examined against laboratory strains by agar well diffusion method and broth dilution assay. Growth kinetics in presence of ß-Lap on Staphylococcus aureus, Staphylococcus epidermidis, and Pseudomonas aeruginosa (ATCC 27853) were assessed by microplate alamarBlue assay. Crystal violet blue assay was used for biofilm inhibition and biofilm eradication. P. aeruginosa catalase (KatA) complexed with ß-Lap was modeled using molecular docking approach. RESULTS: ß-Lap exhibited potent antimicrobial activity against laboratory strains of bacteria with MIC of 0.2 mM for S. saprophyticus and Staphylococcus aureus, and 0.04 mM for Staphylococcus epidermidis and Pseudomonas aeruginosa ATCC 27853. The inhibition of catalase enzyme was found to be the cause for its antibacterial activity. Bioinformatics analysis suggests that ß-Lap can inhibit KatA activity by interacting with catalase proximal active site and heme binding site. The activity of some commercial antibiotics was enhanced in association with ß-Lap. In addition, ß-Lap inhibited the biofilm formation and eradicated the already formed and ultra-mature biofilms of aforesaid bacterial strains. CONCLUSION: These observations indicated that ß-Lap could be a promising antibacterial agent for the treatment and prevention of infectious diseases.

Therapeutic approaches for chronic hepatitis C: a concise review.

Hepatitis C virus (HCV) infection is a significant global health concern, prompting the need for effective treatment strategies. This in-depth review critically assesses the landscape of HCV treatment, drawing parallels between traditional interferon/ribavirin therapy historically pivotal in HCV management and herbal approaches rooted in traditional and complementary medicine. Advancements in therapeutic development and enhanced clinical outcomes axis on a comprehensive understanding of the diverse HCV genome, its natural variations, pathogenesis, and the impact of dietary, social, environmental, and economic factors. A thorough analysis was conducted through reputable sources such as Science Direct, PubMed, Scopus, Web of Science, books, and dissertations. This review primarily focuses on the intricate nature of HCV genomes and explores the potential of botanical drugs in both preventing and treating HCV infections.

Artificial intelligence and cheminformatics tools: a contribution to the drug development and chemical science.

In the ever-evolving field of drug discovery, the integration of Artificial Intelligence (AI) and Machine Learning (ML) with cheminformatics has proven to be a powerful combination. Cheminformatics, which combines the principles of computer science and chemistry, is used to extract chemical information and search compound databases, while the application of AI and ML allows for the identification of potential hit compounds, optimization of synthesis routes, and prediction of drug efficacy and toxicity. This collaborative approach has led to the discovery, preclinical evaluations and approval of over 70 drugs in recent years. To aid researchers in the pursuit of new drugs, this article presents a comprehensive list of databases, datasets, predictive and generative models, scoring functions and web platforms that have been launched between 2021 and 2022. These resources provide a wealth of information and tools for computer-assisted drug development, and are a valuable asset for those working in the field of cheminformatics. Overall, the integration of AI, ML and cheminformatics has greatly advanced the drug discovery process and continues to hold great potential for the future. As new resources and technologies become available, we can expect to see even more groundbreaking discoveries and advancements in these fields.Communicated by Ramaswamy H. Sarma.

Prevalence of Clostridium Difficile Infection (CDI) among Inflammatory Bowel Disease (IBD) Patients in Comparison to Non-IBD Patients in King Abdulaziz Medical City in Jeddah.

Background: The prevalence of Clostridium difficile infection (CDI) as a common complication among inflammatory bowel disease (IBD) has been reported to increase worldwide and has been associated with a poor IBD outcome. Objectives: In this study, our aim was to report on the prevalence of CDI among IBD vs. non-IBD patients in King Abdulaziz Medical City (KAMC). Methods: This retrospective descriptive study was carried out between 2016 and 2020. Data of 89 patients reported with CDI in KAMC were analyzed for demographics and correlations between various characteristics such as BMI, personal/family history of IBD, infection with CDI, diagnosis, method of diagnosis, and treatment modalities. Results: Of the total 89 CDI patients, 59 (66.3%) were adults and 30 (33.7%) were pediatric, of which 36 (40.4%) were females and 53 (59.6%) were males. PCR was the main method of choice for the diagnosis of CDI (89.9%) followed by a positive-culture result (10.0%). Seventy-eight (87.6%) CDI patients were found to be immunocompromised, with two patients diagnosed with IBDs, one with UC, and one with CD. The recurrence rate was 38.4 (30 patients) among the immunocompromised group in comparison to 27.2 (3 patients) in the immunocompetent group (p=0.584). Conclusion: In this study, we found that adults were more prone to CDI infection, especially within hospital settings, and most of the CDI infections occurred in immunocompromised individuals, with cancer as the most common cause of it.

Design-of-Experiment-Assisted Fabrication of Biodegradable Polymeric Nanoparticles: In Vitro Characterization, Biological Activity, and In Vivo Assessment.

Berberine (BER) is an alkaloid obtained from berberis plant having broad biological activities including anticancer. BER-encapsulated alginate (ALG)/chitosan (CHS) nanoparticles (BER-ALG/CHS-NPs) were developed for long-acting improved treatment in breast cancer. The surface of the NPs was activated by a conjugation reaction, and thereafter, the BER-ALG/CHS-NP surface was grafted with folic acid (BER-ALG/CHS-NPs-F) for specific targeting in breast cancer. BER-ALG/CHS-NPs-F was optimized by applying the Box-Behnken design using Expert design software. Moreover, formulations are extensively evaluated in vitro for biopharmaceutical performances and tested for cell viability, cellular uptake, and antioxidant activity. The comparative pharmacokinetic study of formulation and free BER was carried out in animals for estimation of bioavailability. The particle size recorded for the diluted sample using a Malvern Zetasizer was 240 ± 5.6 nm. The ζ-potential and the predicted % entrapment efficiency versus (vs) observed were +18 mV and 83.25 ± 2.3% vs 85 ± 3.5%. The high % drug release from the NPs was recorded. The analytical studies executed using infrared spectroscopy, differential scanning calorimetry, and X-ray diffraction expressed safe combinations of the components in the formulation and physical state of the drug revealed to be amorphous in the formulation. Cytotoxicity testing demonstrated that the formulation effectively lowered the cell viability and IC50 of the tested cell line in comparison to a raw drug. The cellular uptake of BER-ALG/CHS-NPs-F was 5.5-fold higher than that of BER-suspension. The antioxidant capacities of BER-ALG/CHS-NPs-F vs BER-suspension by the DPPH assay were measured to be 62.3 ± 2.5% vs 30 ± 6%, indicating good radical scavenging power of folate-conjugated NPs. The developed formulation showed a 4.4-fold improved oral bioavailability compared to BER-suspension. The hemolytic assay intimated <2% destruction of erythrocytes by the developed formulation. The observed experimental characterization results such as cytotoxicity, cellular uptake, antioxidant activity, and improved absorption suggested the effectiveness of BER-ALG/CHS-NPs-F toward breast cancer.

Antibiotics resistance as a major public health concern: A pharmaco-epidemiological study to evaluate prevalence and antibiotics susceptibility-resistance pattern of bacterial isolates from multiple teaching hospitals.

BACKGROUND: Multi Drugs Resistance (MDR) is among the most worrisome healthcare issues resulting from inappropriate and indiscriminate utilization of antimicrobial agents which has compromised the efficacy and reliability of antimicrobial agents (AMAs). This has not only put a huge burden on the health care system but also is a major cause of morbidity and mortality. This project was designed to evaluate the prevalence of various microbial strains among patients admitted to various teaching hospitals and to assess their susceptibility and resistance towards clinically approved antibiotics. METHODS: The study was conducted during August 2021-February 2022 to determine the prevalence of common resistant strains of bacteria and to analyze their susceptibility pattern to the commonly prescribed antibiotics using standard procedures. One hundred and thirty biological samples including urine, blood, cerebrospinal fluid (CSF), wound swabs, pus and sputum were collected from the site of infection from the patients admitted at different wards of North West General Hospital (NWGH), Peshawar, Pakistan, Khyber Teaching Hospital (KTH), Peshawar Pakistan, and Hayat Abad Medical Complex (HMC) Peshawar Pakistan. Samples were collected and cultured following standard hospital procedures. The cultured samples were subjected to identification procedures including Gram staining, morphological characterization of bacterial colonies and biochemical assessments. The identified bacteria were tested for their susceptibility using Kirby-Bauer disc diffusion method. The diameter of Inhibitory Zones (DIZ) was analyzed following Clinical and Laboratory Standards Institute (CLSI) criteria. Minimum Inhibitory Concentrations (MICs) were evaluated using agar dilution method. Antimicrobials sensitivity were presented as antibiogram following CLSI M39 standard. RESULTS: A total of one hundred and thirty biological samples were collected, out of which one hundred and nine samples were positive for bacterial growth and were further processed for detailed analysis. The frequency and type of bacteria isolated from various cultures indicated that Gram negative bacteria (n = 92/109) were more dominant than Gram-positive (n = 17/109) pathogens. The most prevalent bacteria isolated was Escherichia coli (29.35 %), followed by Staphylococcus aureus (15.59 %), and Klebsiella spp, (12.84 %). In addition, other pathogens including, Enterobacter spp, Citrobacter spp, and Acinetobacter spp. showed a prevalence of 9.175 %, 8.25 %, and 5.50 % respectively. As indicated in the antbiogram, several organisms exhibited considerble decline in the sensitivies towards various antibiotics. A high percentage of resistance was observed against some antibiotics including trimethoprim, co-trimoxazole, amoxicillin/clavulanate, ciprofloxacin, piperacillin/tazobactam, cefotaxime and ceftazidime. CONCLUSION: The prevalence of resistant strains of pathogens is increasing day by day, while the antibiotics commonly prescribed against them are losing their efficacy, which is pushing the world to the era of pre-antibiotics. Unfortunately, the discovery of novel antibiotics is limited and researchers speculate that the is pushing towards pre-antibiotics era. Subsequently, efforts must be directed towards ensuring rational antibiotics use to prevent emergence of MDR pathogens. Our findings indicated that Gram negative bacteria including Escherichia coli was most prevalent. Other bacterial strains including S. aureus, Klebsiella spp, Enterobacter spp, Citrobacter spp, and Acinetobacter spp. were found among the causative agents. Unfortunately, considerable decline in the sensitivities of various bacterial isolated were observed towards the tested antibiotics. Previous studies reported the high prevalence of E. coli and S. aureus in clinical samples of Pakistani hospitals including hospitals in Peshawar and thus our findings are in agreement with the previous reports. Pharmacists being experts can play their role by promoting the optimal use of antimicrobial agents and educating healthcare professionals, patients and the public.

BV6 enhances apoptosis in Lung cancer cells by ameliorating caspase expressions through attenuation of XIAP, cIAP-1, and cIAP-2 proteins.

Objective: The present study aimed to investigate the inhibitory role of second mitochondria determined activator of caspases mimetic on inhibitor of apoptosis proteins (IAPs) and regulation of caspases in nonsmall cell lung cancer cell line. Materials and Methods: Dimethyl sulfoxide and 3-(4, 5-dimethyl thizol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay was done to determine the IC50 of BV6 using NCI-H23 cell line. The levels of mRNA of X-linked IAP (XIAP), cellular IAP (cIAP-1), cIAP-2, caspase-6, and caspase-7 in H23 cell line were evaluated by a quantitative real-time polymerase chain reaction, while their protein expressions were tested using western blotting. Results: Two doses of BV6 dependently downregulated the expression of mRNA of XIAP (P = 0.002, P= 0.0003 vs. untreated), cIAP-1 (P = 0.05, P = 0.005 vs. untreated), and cIAP-2 (P = 0.001, P = 0.0002 vs. untreated), respectively, while the compound upregulated the mRNA expression of caspase-6 (P = 0.001, P < 0.0001 vs. untreated) and caspase-7 (P = 0.001, P = 0.0004 vs. untreated), respectively. Dose dependent of BV6 treatment significantly decreased the protein level of XIAP (P = 0.003, P = 0.007 vs. untreated), cIAP-1 (P = 0.02, P = 0.01 vs. untreated), and cIAP-2 (P = 0.008,P = 0.008 vs. untreated), respectively. However, the compound increased the protein level of caspase-6 and caspase-7 when compared to untreated control (P = 0.006,P = 0.001) and (P = 0.01, P = 0.001), respectively. Conclusions: The result showed that BV6 treatment reduced the level of mRNA of XIAP, cIAP-1, and cIAP-2 and increased the gene expression of caspase-6 and caspase-7 in NCI-H23 cell line. Therefore, the study revealed that BV6 could be used in future as additional therapeutics in lung cancer.

Phytochemical Screening and In-Vitro Antibacterial and Anticancer Activity of Crude Extract of Matricaria aurea.

BACKGROUND: Infectious diseases constantly represent the source of sickness as well as mortality in human beings. Herbal applications in human life through using plants for antibacterial and anticancer activity have shown the potential medicinal outcome. OBJECTIVES: To evaluate the antibacterial and anticancer activities of the crude extract of Matricaria aurea. MATERIALS AND METHODS: The antibacterial activity of the crude flowers of M. aurea extract was examined against reference and clinical bacterial strains by agar well diffusion method. Minimum inhibitory concentrations and minimum bactericidal concentrations were determined by micro broth dilution assays using MH broth. Herbal extract was employed over human breast adenocarcinoma cell line (MCF-7), hepatocellular carcinoma cell line (HepG-2) and colorectal adenocarcinoma cell line (HCT-116) to optimize cancer cells proliferation by SRB assay. RESULTS: The data has shown that the extract from M. aurea had significant antimicrobial activity against the tested microorganisms. The plant extract showed higher antibacterial activity against the reference strain of Streptococcus pyogenes. The MIC and MBC varied between 0.38-12.5 mg/ml and 3.1-200 mg/ml respectively. Synergy study elucidated the significant bacteriostatic effect of M. aurea extract on S. aureus and S. saprophyticus. The data of SRB assay deliver the potential anticancer activity through cell death. CONCLUSION: This study delivers innovative information that M. aurea possessed excellent bio-activities against pathogenic microbes and cancer cells, which drive attention for further research to explore the active components responsible for biological efficacies.

MtFtsX a predicted membrane domain of ABC transporter complex MtFtsEX of Mycobacterium tuberculosis interacts with the cell division protein MtFtsZ.

Background: Bacterial cytokinesis is orchestrated by a complex of dozen of proteins called 'divisome' at the mid-cell site. FtsZ, the eukaryotic tubulin homolog, localizes to the mid-cell site where it polymerizes and forms a cytokinetic Z-ring. The Z-ring acts as a docking platform for other proteins to localize. In model organisms, Escherichia coli and Bacillus subtilis, FtsZ is known to interact with several proteins. The role of few of these interactions is known, while of others is yet to be studied. In Mycobacterium tuberculosis, the cell division and its regulation are poorly studied. Although, most of the divisome proteins are conserved in M. tuberculosis, surprisingly the homologues of the protein factors required for membrane association of Z-ring and its stabilization are absent. In E. coli FtsE and FtsX, the constituent ATPase and membrane domains of the ABC transporter complex, localize to the Z-ring immediately after Z-ring stabilizing proteins, ZipA and FtsA. Therefore, investigation of the interaction between MtFtsX and MtFtsZ is demanding. Methods: Bacterial two-hybrid system was used to identify the interaction between MtFtsE and MtFtsZ. This interaction was further confirmed by biochemical methods of Ni2+-NTA agarose pull-down and coimmunoprecipitation. Results and Conclusion: Here, we demonstrated that MtFtsX interacts with MtFtsZ in vivo and ex-vivo. Further, we showed that self-interacting MtFtsX interacts with MtFtsE. However, we did not find any interaction between MtFtsE and MtFtsZ. These results suggest that the membrane domain MtFtsX of the ABC transporter complex 'MtFtsEX' might be the membrane-tethering and stabilizing factor for Z-ring in M. tuberculosis.

Mycobacterium tuberculosis cell division protein, FtsE, is an ATPase in dimeric form.

FtsE is one of the earliest cell division proteins that assembles along with FtsX at the mid-cell site during cell division in Escherichia coli. Both these proteins are highly conserved across diverse bacterial genera and are predicted to constitute an ABC transporter type complex, in which FtsE is predicted to bind ATP and hydrolyse it, and FtsX is predicted to be an integral membrane protein. We had earlier reported that the MtFtsE of the human pathogen, Mycobacterium tuberculosis, binds ATP and interacts with MtFtsX on the cell membrane of M. tuberculosis and E. coli. In this study, we demonstrate that MtFtsE is an ATPase, the active form of which is a dimer, wherein the participating monomers are held together by non-covalent interactions, with the Cys84 of each monomer present at the dimer interface. Under oxidising environment, the dimer gets stabilised by the formation of Cys84-Cys84 disulphide bond. While the recombinant MtFtsE forms a dimer on the membrane of E. coli, the native MtFtsE seems to be in a different conformation in the M. tuberculosis membrane. Although disulphide bridges were not observed on the cytoplasmic side (reducing environment) of the membrane, the two participating monomers could be isolated as dimers held together by non-covalent interactions. Taken together, these findings show that MtFtsE is an ATPase in the non-covalent dimer form, with the Cys84 of each monomer present in the reduced form at the dimer interface, without participating in the dimerisation or the catalytic activity of the protein.

Identification and semi-quantitative analysis of Mycobacterium tuberculosis H37Rv ftsZ gene-specific promoter activity-containing regions.

The cytokinetic protein FtsZ plays a pivotal role in regulation of cell division in bacteria. Multiple promoters regulate transcription of the ftsZ gene in Escherichia coli, Streptomyces and Bacillus species. In order to identify promoter activity-containing regions of the ftsZ gene of Mycobacterium tuberculosis H37Rv (MtftsZ) in vivo, different regions upstream of MtftsZ, namely, the ftsQ-ftsZ intergenic region, the ftsQ open reading frame (ORF), and different regions of ftsQ ORF, were analyzed in a gfp reporter plasmid in Mycobacterium smegmatis mc(2)155 cells. Flow cytometric analysis of mid-logarithmic M. smegmatis mc(2)155 cells containing these transcription fusion constructs revealed GFP expression in the cells harboring the ftsQ-ftsZ intergenic region (172 bp), the entire ftsQ ORF (945 bp), and 5' 467-bp and 3' 217-bp regions of ftsQ ORF. RT-PCR analyses on RNA from M. smegmatis mc(2)155 cells, transformed with the entire ftsQ ORF-ftsQ-ftsZ intergenic region-containing construct, as well as on RNA from M. tuberculosis, confirmed that the regions identified indeed elicit promoter activity. Semi-quantitative RT-PCR analyses of gfp transcripts driven by cloned MtftsZ promoter regions in M. smegmatis cells showed threefold higher promoter activity from ftsQ ORF than from the ftsQ-ftsZ intergenic region. Expression from the individual 5' and 3' regions of ftsQ ORF was almost equivalent to that from the ftsQ-ftsZ intergenic region. RT-PCR analyses on RNA from M. tuberculosis quantitatively confirmed these promoter activities. Thus, at least three independent regions in the immediate upstream sequence of MtftsZ contain promoter activity, with the major contribution coming from ftsQ ORF.

Molecular characterisation of ABC transporter type FtsE and FtsX proteins of Mycobacterium tuberculosis.

Elicitation of drug resistance and various survival strategies inside host macrophages have been the hallmarks of Mycobacterium tuberculosis as a successful pathogen. ATP Binding Cassette (ABC) transporter type proteins are known to be involved in the efflux of drugs in bacterial and mammalian systems. FtsE, an ABC transporter type protein, in association with the integral membrane protein FtsX, is involved in the assembly of potassium ion transport proteins and probably of cell division proteins as well, both of which being relevant to tubercle bacillus. In this study, we cloned ftsE gene of M. tuberculosis, overexpressed and purified. The recombinant MtFtsE-6xHis protein and the native MtFtsE protein were found localized on the membrane of E. coli and M. tuberculosis cells, respectively. MtFtsE-6xHis protein showed ATP binding in vitro, for which the K42 residue in the Walker A motif was found essential. While MtFtsE-6xHis protein could partially complement growth defect of E. coli ftsE temperature-sensitive strain MFT1181, co-expression of MtFtsE and MtFtsX efficiently complemented the growth defect, indicating that the MtFtsE and MtFtsX proteins might be performing an associated function. MtFtsE and MtFtsX-6xHis proteins were found to exist as a complex on the membrane of E. coli cells co-expressing the two proteins.