Mycobacterium Tuberculosis Protein Research Articles

The SH3-binding domain of chorismate mutase protein of Mycobacterium tuberculosis contributes to mycobacterial virulence

Crystal structure of the secretory chorismate mutase protein of Mycobacterium tuberculosis (MtbCM) reveals presence of a proline rich region on its surface that serve as a recognition site for protein-protein interaction. This study shows that MtbCM upregulates IL-10 which favors M.tuberculosis by affecting PKCε-MKP-1-p38 MAPK signaling. MtbCM translocates to the Golgi-network where it interacts with AKAP9 via its SH3-binding domain to inhibit AKAP9-PKCε interaction and reducing PKCε phosphorylation. In the absence of phosphorylated PKCε, IRAK3 fails to stabilize MKP-1 resulting in higher p38 MAPK activation and IL-10 production. M. smegmatis expressing MtbCM survived better in infected mice. Mutation in SH3-binding domain ablated MtbCM-AKAP9 interaction resulting in IL-10 production and decreased bacterial survival. This study highlights the importance of SH3-binding domain in host-pathogen interaction and a role of MtbCM in modulation of cytokine response and mycobacterial virulence in addition to its role in shikimate pathway.

Mycobacterial peptidyl prolyl isomerase A activates STING-TBK1-IRF3 signaling to promote IFNβ release in macrophages.

Peptidyl prolyl isomerases (PPIases) are well-conserved protein-folding enzymes that moonlight as regulators of bacterial virulence. Peptidyl prolyl isomerase A, PPiA (Rv0009) is a secretory protein of Mycobacterium tuberculosis that possesses sequence and structural similarity to eukaryotic cyclophilins. In this study, we validated the interaction of PPiA with stimulator of interferon genes (STING) using both, Escherichia coli-based and mammalian in vitro expression systems. In vitro pull-down assays confirmed that the cytosolic domain of STING interacts with PPiA, and moreover, we found that PPiA could induce dimerization of STING in macrophages. In silico docking analyses suggested that the PXXP (PDP) motif of PPiA is crucial for interaction with STING, and concordantly, mutations in the PDP domain (PPiA MUT-II) abrogated this interaction, as well as the ability of PPiA to facilitate STING dimerization. In agreement with these observations, fluorescence microscopy demonstrated that STING and wild-type PPiA, but not PPiA MUT-II, could colocalize when expressed in HEK293 cells. Highlighting the importance of the PDP domain further, PPiA, but not PPiA MUT-II could activate Tank binding kinase 1 (TBK1)-interferon regulatory factor 3 (IRF3) signaling to promote the release of interferon-beta (IFNβ). PPiA, but not PPiA MUT-II expressed in Mycobacterium smegmatis induced IFNβ release and facilitated bacterial survival in macrophages in a STING-dependent manner. The PPiA-induced release of IFNβ was c-GAS independent. We conclude that PPiA is a previously undescribed mycobacterial regulator of STING-dependent type I interferon production from macrophages.

Design of molecular inhibitors against the NuoC protein of Mycobacterium tuberculosis.

Tuberculosis (TB) is one of the main health issues for the international scientific community. The NADH- quinone oxidoreductase subunit C (NuoC) protein belongs to the NADH dehydrogenase family, which plays a critical role in the electron transport chain and ATP synthesis and energy production in Mycobacterium tuberculosis (Mtb). In the present study, Nuoc protein is considered a potential drug target for identifying inhibitors for the protein. The Nuoc protein’s 3D structural features are determined using computational techniques, including active site identification, ADME properties and virtual screening. The NuoC theoretical model was developed using homology modeling techniques and its structure was verified by various validation methods. They provide insight into the conformational changes occurring in the NuoC protein. To identify drug-like compounds virtual screening studies were conducted around the active site using several ligand databases. According to the research residues the amino acid residues ARG98, ARG75 (basic), ASP99, ASP189, ASP98 (acidic), LEU101, LEU194 (nonpolar neutral), THR180 (polar neutral), GLU177(polar neutral), TYR181(polar neutral), PRO102, PRO192 (nonpolar neutral), and HIS191(basic) are important in drug-target interactions. Additionally, docking experiments of TB medications against NuoC were carried out. The study found that the ADME parameters of the selected ligand molecules are more favourable compared to existing TB drugs. This highlights the drug-like activity of the ligand molecules in inhibiting NuoC proteins. The structural data, along with the information about the active site and the selected ligand molecules, can help in identifying new therapeutic scaffolds for the treatment of Tuberculosis.

Recombinant mycobacterial DNA-binding protein 1 with post-translational modifications boosts IFN-gamma production from BCG-vaccinated individuals’ blood cells in combination with CpG-DNA

Tuberculosis remains a large health threat, despite the availability of the tuberculosis vaccine, BCG. As BCG efficacy gradually decreases from adolescence, BCG-Prime and antigen-booster may be an efficient strategy to confer vaccine efficacy. Mycobacterial DNA-binding protein 1 (MDP1, namely Rv2986c, hupB or HU) is a major Mycobacterium tuberculosis protein that induces vaccine-efficacy by co-administration with CpG DNA. To produce MDP1 for booster-vaccine use, we have created recombinant MDP1 produced in both Escherichia coli (eMDP1) and Mycolicibacterium smegmatis (mMDP1), an avirulent rapid-growing mycobacteria. We tested their immunogenicity by checking interferon (IFN)-gamma production by stimulated peripheral blood cells derived from BCG-vaccinated individuals. Similar to native M. tuberculosis MDP1, we observed that most lysin resides in the C-terminal half of mMDP1 are highly methylated. In contrast, eMDP1 had less post-translational modifications and IFN-gamma stimulation. mMDP1 stimulated the highest amount of IFN-gamma production among the examined native M. tuberculosis proteins including immunodominant MPT32 and Antigen 85 complex. MDP1-mediated IFN-gamma production was more strongly enhanced when combined with a new type of CpG DNA G9.1 than any other tested CpG DNAs. Taken together, these results suggest that the combination of mMDP1 and G9.1 possess high potential use for human booster vaccine against tuberculosis.

Cyclic AMP binding to a universal stress protein in Mycobacterium tuberculosis is essential for viability

Mycobacterial genomes encode multiple adenylyl cyclases and cAMP effector proteins, underscoring the diverse ways these bacteria utilize cAMP. We identified universal stress proteins, Rv1636 and MSMEG_3811 in Mycobacterium tuberculosis andMycobacterium smegmatis, respectively, as abundantly expressed, novel cAMP-binding proteins. Rv1636 is secreted via the SecA2 secretion system in M.tuberculosis but is not directly responsible for the efflux of cAMP from the cell. In slow-growing mycobacteria, intrabacterial concentrations of Rv1636 were equivalent to the concentrations of cAMP present in the cell. In contrast, levels of intrabacterial MSMEG_3811 in M.smegmatis were lower than that of cAMP and therefore, overexpression of Rv1636 increased levels of "bound" cAMP. While msmeg_3811 could be readily deleted from the genome of M.smegmatis, we found that the rv1636 gene is essential for the viability of M.tuberculosis and is dependent on the cAMP-binding ability of Rv1636. Therefore, Rv1636 may function to regulate cAMP signaling by direct sequestration of the second messenger. This is the first evidence of a "sponge" for any second messenger in bacterial signaling that would allow mycobacterial cells to regulate the available intrabacterial "free" pool of cAMP.

The role of transcriptional regulators in metal ion homeostasis of Mycobacterium tuberculosis.

Metal ions are essential trace elements for all living organisms and play critical catalytic, structural, and allosteric roles in many enzymes and transcription factors. Mycobacterium tuberculosis (MTB), as an intracellular pathogen, is usually found in host macrophages, where the bacterium can survive and replicate. One of the reasons why Tuberculosis (TB) is so difficult to eradicate is the continuous adaptation of its pathogen. It is capable of adapting to a wide range of harsh environmental stresses, including metal ion toxicity in the host macrophages. Altering the concentration of metal ions is the common host strategy to limit MTB replication and persistence. This review mainly focuses on transcriptional regulatory proteins in MTB that are involved in the regulation of metal ions such as iron, copper and zinc. The aim is to offer novel insights and strategies for screening targets for TB treatment, as well as for the development and design of new therapeutic interventions.

A novel enzyme-linked ligand-sorbent assay (ELLSA) to screening pulmonary tuberculosis: a retrospective cross-sectional study

BackgroundLittle knowledge of antigen existence in the pulmonary tuberculosis (PTB) patient serum impeded its development in antigen detection technology, despite its considerable potential. MethodsHuman ligand proteins and their adsorbent Mycobacterium tuberculosis (M.tb) proteins in the serum of PTB patients were identified using human protein chip (HuProt™) and LC-MS/MS, successively. The monoclonal antibody of ligand proteins, C5orf24, and polyclonal antibody of 9 M.tb proteins were prepared on mice and rabbits which were used to develop a novel enzyme-linked ligand-sorbent assay (ELLSA). The 412 volunteers were divided into the PTB group (n = 250) and the healthy control (n = 162). The PTB group was further divided into ATB (n = 131), LTBI (n = 18), Clinical diagnosis (n = 18), and Suspected (n = 73). All samples were tested by ELLSA to evaluate the diagnostic performance of ELLSA in PTB patients. ResultsNine ligand proteins specific to PTB patients were identified on chips, with Chromosome 5 Open Reading Frame 24 (C5orf24) and kinocilin (KNCN) showing significantly higher signals. Proteomic analysis of the C5orf24-and KNCN-adsorbent protein complexes revealed 10 and 10 of the M.tb proteins, respectively. According to the composition reference of standard, the ELLSA based on C5orf24 ligand demonstrated a higher sensitivity of 69.6% and specificity of 90.18% in ATB patients and had a sensitivity of 64.22% in bacterial negative pulmonary tuberculosis, whereas the sensitivity of MGIT 960 and Xpert M.tb/RIF were 0%, respectively. ConclusionsM.tb proteins in serum can be enriched by ligand proteins and detected by ELLSA which proved to have excellent diagnostic performance for PTB.

Elucidating the network interactions between 21 secreted Mycobacterium tuberculosis proteins and host proteins: the role of DnaK in enhancing Mtb survival via LDHB.

Elucidating the network interactions between 21 secreted Mycobacterium tuberculosis proteins and host proteins: the role of DnaK in enhancing Mtb survival via LDHB.

Proteomic Analysis of the Mycobacterium tuberculosis Outer Membrane for Potential Implications in Uptake of Small Molecules.

Increased resistance to current antimycobacterial agents and a potential bias toward relatively hydrophobic chemical entities highlight an urgent need to understand how current anti-TB drugs enter the tubercle bacilli. While inner membrane proteins are well-studied, how small molecules cross the impenetrable outer membrane remains unknown. Here, we employed mass spectrometry-based proteomics to show that octyl-β-d-glucopyranoside selectively extracts the outer membrane proteins of Mycobacterium tuberculosis. Differentially expressed proteins between nutrient-replete and nutrient-depleted conditions were enriched to identify proteins involved in nutrient uptake. We demonstrate cell surface localization of seven new proteins using immunofluorescence and show that overexpression of the proteins LpqY and ProX leads to hypersensitivity toward streptomycin, while overexpression of SubI, SpmT, and Rv2041 exhibited higher membrane permeability, assessed through an EtBr accumulation assay. Further, proton NMR metabolomics suggests the role of six outer membrane proteins in glycerol uptake. This study identifies several outer membrane proteins that are involved in the permeation of small hydrophilic molecules and are potential targets for enhancing the uptake and efficacy of anti-TB drugs.

Mycobacterium tuberculosis protein PPE15 (Rv1039c) possesses eukaryote-like SH3 domain that interferes with NADPH Oxidase assembly and Reactive Oxygen Species production

Inhibition of Reactive Oxygen Species (ROS) is one of the strategies that Mycobacterium tuberculosis (Mtb) employs as its defence mechanism. In this study, the role of PPE15 (Rv1039c), a late-stage protein, has been investigated in modulating the cellular ROS. We discovered PPE15 to be a secretory protein that downregulates ROS generation in THP1 macrophages. Our in-silico analysis revealed the presence of a eukaryote-like SH3 (SH3e) domain in PPE15. The predicted SH3e-domain of PPE15 was found to interact with cytosolic components of NADPH Oxidase (NOX), p67phox and p47phox through molecular docking. In-vitro experiments using THP1 macrophages showed a diminished NADP/NADPH ratio, indicating reduced NOX activity. We also observed increased levels of p67phox and p47phox in the cytoplasmic fraction of PPE15 treated macrophages as compared to the plasma membrane fraction. To understand the role of the SH3e-domain in ROS modulation, this domain was deleted from the full-length PPE15 (PPE15-/-SH3). We observed an increase in cellular ROS and NADP/NADPH ratio in response to PPE15-/-SH3 protein. The interaction of PPE15-/-SH3 with p67phox or p47phox was also reduced in the cytoplasm, indicating migration of NOX subunits to the plasma membrane. Additionally, M. smegmatis expressing PPE15 was observed to be resistant to oxidative stress with significant intracellular survival in THP1 macrophages as compared to M. smegmatis expressing PPE15-/-SH3. These observations suggest that the SH3e-domain of PPE15 interferes with ROS generation by sequestering NOX components that inhibit NOX assembly at the cell membrane. Therefore, PPE15 acts like a molecular mimic of SH3-domain carrying eukaryotic proteins that can be employed by Mtb at late stages of infection for its survival. These findings give us new insights about the pathogen evading strategy of Mtb which may help in improving the therapeutics for TB treatment.

Metabolites from Streptomyces aureus (VTCC43181) and Their Inhibition of Mycobacterium tuberculosis ClpC1 Protein.

Tuberculosis is one of the most common infectious diseases in the world, caused by Mycobacterium tuberculosis. The outbreak of multiple drug-resistant tuberculosis has become a major challenge to prevent this disease worldwide. ClpC1 is a Clp ATPase protein of Mycobacterium tuberculosis, functioning as a chaperon when combined with the Clp complex. ClpC1 has emerged as a new target to discover anti-tuberculosis drugs. This study aimed to explore the ClpC1 inhibitors from actinomycetes, which have been known to provide abundant sources of antibiotics. Two cyclic peptides, including nocardamin (1), halolitoralin A (3), and a lactone pleurone (2), were isolated from the culture of Streptomyces aureus (VTCC43181). The structures of these compounds were determined based on the detailed analysis of their spectral data and comparison with references. This is the first time these compounds have been isolated from S. aureus. Compounds 1-3 were evaluated for their affection of ATPase activity of the recombinant ClpC1 protein. Of these compounds, halolitoralin A (1), a macrocyclic peptide, was effective for the ATPase hydrolysis of the ClpC1 protein.

Development of NP-Based Universal Vaccine for Influenza A Viruses.

The nucleoprotein (NP) is a vital target for the heterosubtypic immunity of CD8+ cytotoxic T lymphocytes (CTLs) due to its conservation among influenza virus subtypes. To further enhance the T cell immunity of NP, autophagy-inducing peptide C5 (AIP-C5) from the CFP10 protein of Mycobacterium tuberculosis was used. Mice were immunized intranasally (i.n.) with human adenoviral vectors, HAd-C5-NP(H7N9) or HAd-NP(H7N9), expressing NP of an H7N9 influenza virus with or without the AIP-C5, respectively. Both vaccines developed similar levels of NP-specific systemic and mucosal antibody titers; however, there was a significantly higher number of NP-specific CD8 T cells secreting interferon-gamma (IFN-γ) in the HAd-C5-NP(H7N9) group than in the HAd-NP(H7N9) group. The HAd-C5-NP(H7N9) vaccine provided better protection following the challenge with A/Puerto Rico/8/1934(H1N1), A/Hong Kong/1/68(H3N2), A/chukkar/MN/14951-7/1998(H5N2), A/goose/Nebraska/17097/2011(H7N9), or A/Hong Kong/1073/1999(H9N2) influenza viruses compared to the HAd-NP(H7N9) group. The autophagy transcriptomic gene analysis of the HAd-C5-NP(H7N9) group revealed the upregulation of some genes involved in the positive regulation of the autophagy process. The results support further exploring the use of NP and AIP-C5 for developing a universal influenza vaccine for pandemic preparedness.

CLPC1 protein inhibition, antimycobacterial, and anti‐inflammatory properties of the metabolite from Streptomyces wuyuanensis collected in Nghe An province, Vietnam

AbstractStreptomyces wuyuanensis A79 was isolated from the soil samples collected in Quynh Luong mangrove forest, Quynh Luu district, Nghe An province, Vietnam. This strain demonstrated antimycobacterial activity against Mycobacterium smegmatis. Compound 1 (indole‐3‐carboxylic acid) was isolated from the culture extract of S. wuyuanensis (A79). This compound displayed the affection of ATPase activity of the recombinant ClpC1 protein, a regulatory protein of Mycobacterium tuberculosis. In addition, compound 1 exhibited anti‐inflammatory activity through the inhibition of TNF‐α on LPS‐stimulated macrophage RAW264.7. This is the first time compound 1 (indole‐3‐carboxylic acid) has been isolated from S. wuyuanensis. Indole‐3‐carboxylic acid was also reported for the first time its inhibition of M. tuberculosis ClpC1 protein.

Devising Isolation Forest-Based Method to Investigate the sRNAome of Mycobacterium tuberculosis Using sRNA-seq Data.

Small non-coding RNAs (sRNAs) regulate the synthesis of virulence factors and other pathogenic traits, which enables the bacteria to survive and proliferate after host infection. While high-throughput sequencing data have proved useful in identifying sRNAs from the intergenic regions (IGRs) of the genome, it remains a challenge to present a complete genome-wide map of the expression of the sRNAs. Moreover, existing methodologies necessitate multiple dependencies for executing their algorithm and also lack a targeted approach for the de novo sRNA identification. We developed an Isolation Forest algorithm-based method and the tool Prediction Of sRNAs using Isolation Forest for the de novo identification of sRNAs from available bacterial sRNA-seq data (http://posif.ibab.ac.in/). Using this framework, we predicted 1120 sRNAs and 46 small proteins in Mycobacterium tuberculosis. Besides, we highlight the context-dependent expression of novel sRNAs, their probable synthesis, and their potential relevance in stress response mechanisms manifested by M. tuberculosis.

The potential of several compounds in Peperomia pellucida as a solution to rifampicin resistance in tuberculosis disease

Background: Tuberculosis is a lung disease caused by Mycobacterium tuberculosis. In 2018, the incidence of tuberculosis in Indonesia was 1,017,290. The number of cases was accompanied by an increase in the number of resistances to first-line antibiotics (rifampicin) used due to patient non-compliance as well as the long duration of treatment. The increase in resistance around the world is always increasing every year by about 2%. Therefore, we need another alternative to existing plants in Indonesia that has the potential to be a solution to the problem and overcome resistance to these treatments.Objective: This review was conducted to find out the potential of Peperomia pellucida as an antituberculosis drug that can be a solution to rifampicin resistance in tuberculosis disease. Results: The results of the literature review showed that the Peperomia pellucida or suruhan plants have antibacterial potential against Mycobacterium tuberculosis because it contains a class of alkaloid compounds that function to inhibit deprenyl-phosphoryl-β-D-ribose-2 oxidase (an enzyme responsible for the biosynthesis of arabinogalactan in the cell walls of Mycobacterium tuberculosis), a flavonoid that has activity inhibition of fatty acid synthase II (FAS-II) enzymes, tannins that can precipitate proteins in Mycobacterium tuberculosis, and triterpenoids that have an activation mechanism of intracellular killing cascades in host cells during tuberculosis bacterial infection.Conclusion: Peperomia pellucida or suruhan plants have the potential to be further investigated as a solution to overcome rifampicin resistance in tuberculosis disease.Keywords: Mycobacterium tuberculosis, rifampicin resistance, Peperomia pellucida

Ultrasensitive biosensing platform for Mycobacterium tuberculosis detection based on functionalized graphene devices.

Tuberculosis (TB) has high morbidity as a chronic infectious disease transmitted mainly through the respiratory tract. However, the conventional diagnosis methods for TB are time-consuming and require specialists, making the diagnosis of TB with point-of-care (POC) detection difficult. Here, we developed a graphene-based field-effect transistor (GFET) biosensor for detecting the MPT64 protein of Mycobacterium tuberculosis with high sensitivity as a POC detection platform for TB. For effective conjugation of antibodies, the graphene channels of the GFET were functionalized by immobilizing 1,5-diaminonaphthalene (1,5-DAN) and glutaraldehyde linker molecules onto the graphene surface. The successful immobilization of linker molecules with spatial uniformity on the graphene surface and subsequent antibody conjugation were confirmed by Raman spectroscopy and X-ray photoelectron spectroscopy. The GFET functionalized with MPT64 antibodies showed MPT64 detection with a detection limit of 1fg/mL in real-time, indicating that the GFET biosensor is highly sensitive. Compared to rapid detection tests (RDT) and enzyme-linked immunosorbent assays, the GFET biosensor platform developed in this study showed much higher sensitivity but much smaller dynamic range. Due to its high sensitivity, the GFET biosensor platform can bridge the gap between time-consuming molecular diagnostics and low-sensitivity RDT, potentially aiding in early detection or management of relapses in infectious diseases.

Drug design and in-silico study of 2-alkoxylatedquinoline-3-carbaldehyde compounds: Inhibitors of Mycobacterium tuberculosis

Tuberculosis (TB) caused by Mycobacterium tuberculosis (MTB) is a deadly communicable disease that frequently affects the lungs. Current treatment protocols are bedeviled by extensive drug-resistant (XDR) and the evolution of multidrug-resistant (MDR-TB) strains. Virtual in-silico drug discovery tools were used to investigate thirty-two hypothetical 2-alkoxylatedquinoline-3-carbaldehyde compounds for screening against ten different diseases proteins based on drug-likeness, oral bioavailability, pharmacokinetics, global chemical reactivity and their theoretical binding affinities. Their chemical structures were optimized at the density functional theory (DFT) using Becke's three-parameter exchange functional with Lee–Yang–Parr correlation function (B3LYP) and the triple zeta basis set 6–311 in a vacuum using Gaussian 09 W software. Docking study using Pyrx and Discovery studio. Fourteen compounds; 4 - 6, 12 – 14, 19 – 22 and 27–30 complied with the established drug-likeness rules, however, five compounds 12, 13, 27, 28 and 29 exhibited no significant toxicity. Structural activity relationship revealed that shorter (n < 3) or longer (n > 5) alkyloxyl substituents at position-2 of the quinoline moiety reduces drug-likeness and increases toxicity. Individually, the binding energies obtained were (-8.9 kcal/ mole) against malaria for compound 12 and (-8.2 kcal/mole) against the diabetes for compound 29, both highest for the ten diseases investigated. Mycobacterium Tuberculosis proteins investigated. Molecular dynamics also confirms that 12 and 27 binds very well in the active pocket of Mycobacterium tuberculosis and calculated total free binding energy from MMPBSA is -97.53 ± 2.47 and -58.62 ± 2.94 kJ/mol respectively. The five lead compounds all had binding energies higher than the reference tuberculosis drugs; Isoniazid and Ethambutanol.

Research progress on Mycobacterium tuberculosis acetyltransferase

The protein acetylation of Mycobacterium tuberculosis(MTB) plays an important role in virulence, drug resistance, regulation of metabolism and host anti-tuberculosis immune response. The proteins acetylation of MTB and host protein could be induced by the MTB acetyltransferase, which is related to the occurrence, development and prognosis of tuberculosis (TB). A clear understanding of the function of MTB acetyltransferase and identification of its targeted regulatory protein acetylation modification is critical to elucidate the pathogenic mechanism and drug resistance mechanism of TB, and then this could then provide new targets for the development of anti-tuberculosis drugs. This article systematically reviewed the research progress on MTB acetyltransferase related functions, which will provide a theoretical basis for further research on its mediated protein acetylation modification, further development of new anti-tuberculosis drugs and elucidation of drug resistance mechanism.

PE12 interaction with TLR4 promotes intracellular survival of Mycobacterium tuberculosis by suppressing inflammatory response

Macrophages serve as the primary immune cells responsible for the innate immune defense against Mycobacterium tuberculosis (MTB) infection within the host. Specifically, NLRP3, a member of the NLRs family, plays a significant role in conferring resistance against MTB infection. Conversely, MTB evades innate immune killing by impeding the activation of the NLRP3 inflammasome, although the precise mechanism remains uncertain. In this study, we have identified PE12 (Rv1172c), a member of the PE/PPE family proteins, as an extracellular protein of MTB. PE12 interacts with Toll like receptor 4 (TLR4) in macrophages, forming the PE12-TLR4 complex which subsequently inhibits the transcription and expression of NLRP3. As a result, the transcription and secretion of IL-1β are reduced through the PE12-TLR4-NLRP3-IL-1β immune pathway. In vitro and in vivo experiments using a PE12-deficient strain (H37RvΔPE12) demonstrate a weakening of the suppression of the inflammatory response to MTB infection. Our findings highlight the role of the PE12 protein in not only inhibiting the transcription and release of inflammatory cytokines but also mediating the killing of MTB escape macrophages through TLR4 and inducing lung injury in MTB-infected mice. These results provide evidence that PE12 plays a significant role in the inhibition of the host immune response by MTB.

The heparin-binding hemagglutinin protein of Mycobacterium tuberculosis is a nucleoid-associated protein

Nucleoid-associated proteins (NAPs) regulate multiple cellular processes such as gene expression, virulence, and dormancy throughout bacterial species. NAPs help in the survival and adaptation of Mycobacterium tuberculosis (Mtb) within the host. Fourteen NAPs have been identified in Escherichia coli; however, only seven NAPs are documented in Mtb. Given its complex lifestyle, it is reasonable to assume that Mtb would encode for more NAPs. Using bioinformatics tools and biochemical experiments, we have identified the heparin-binding hemagglutinin (HbhA) protein of Mtb as a novel sequence-independent DNA-binding protein which has previously been characterized as an adhesion molecule required for extrapulmonary dissemination. Deleting the carboxy-terminal domain of HbhA resulted in a complete loss of its DNA-binding activity. Atomic force microscopy showed HbhA-mediated architectural modulations in the DNA, which may play a regulatory role in transcription and genome organization. Our results showed that HbhA colocalizes with the nucleoid region of Mtb. Transcriptomics analyses of a hbhA KO strain revealed that it regulates the expression of ∼36% of total and ∼29% of essential genes. Deletion of hbhA resulted in the upregulation of ∼73% of all differentially expressed genes, belonging to multiple pathways suggesting it to be a global repressor. The results show that HbhA is a nonessential NAP regulating gene expression globally and acting as a plausible transcriptional repressor.