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 Table of Contents  
Year : 2016  |  Volume : 5  |  Issue : 1  |  Page : 83-88

Interaction of antimicrobial peptide with mycolyl transferase in Mycobacterium tuberculosis

Ashok & Rita Patel Institute of Integrated Study & Research in Biotechnology and Allied Sciences, Adit Campus, New Vallabh Vidhyanagar, Gujarat, India

Date of Web Publication8-Feb-2017

Correspondence Address:
Devjani I Banerjee
Ashok & Rita Patel Institute of Integrated Study & Research in Biotechnology and Allied Sciences, Adit Campus, New Vallabh Vidhyanagar, Anand, Gujarat 388121
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Source of Support: None, Conflict of Interest: None

DOI: 10.1016/j.ijmyco.2015.07.002

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It is estimated that about 40% of the Indian population are infected with tuberculosis (TB) and that ~3,000,000 people die as a result of TB annually. TB is caused by Mycobacterium tuberculosis. In 2011, the World Health Organization declared India as having the highest TB burden worldwide. An important criteria for pathogenicity is the presence of mycolic acid linked to the protective outer membrane of bacteria. Mycolyl transferase catalyzes the transfer of mycolic acid and promotes cell wall synthesis. This is also considered as a novel target for drug-mediated intervention strategies. Here, we have attempted to understand the interaction between the antimicrobial peptide (AMP), dermcidin, and mycolyl transferase in M. tuberculosis using a computational approach. The present study was undertaken in order to elucidate the capability of AMPs to treat this bacteria, which is less sensitive to available antibiotics, and to design a novel method for new therapies.

Keywords: Antimicrobial peptides, Dermcidin, Mycobacterium tuberculosis, Mycolyl transferase

How to cite this article:
Banerjee DI, Gohil TP. Interaction of antimicrobial peptide with mycolyl transferase in Mycobacterium tuberculosis. Int J Mycobacteriol 2016;5:83-8

How to cite this URL:
Banerjee DI, Gohil TP. Interaction of antimicrobial peptide with mycolyl transferase in Mycobacterium tuberculosis. Int J Mycobacteriol [serial online] 2016 [cited 2023 Apr 1];5:83-8. Available from: https://www.ijmyco.org/text.asp?2016/5/1/83/199748

  Introduction Top

According to 2013 World Health Organization statistics for Mycobacterium tuberculosis (MTB) in India, approximately 2.1 billion cases have been reported out of global incidences of 9 million TB cases. Such high incidences of TB in India indicate a need for better health habits and food quality to prevent its spread [1]. Initially, TB treatment was possible using conventional antibiotics, however, the scenario has worsened due to the emergence of drug-resistant MTB. The phenomena of drug resistance began with multidrug resistant tuberculosis, followed by the discovery of extensively drug-resistant TB and total drug-resistant TB. Acquisition of resistance to many antibiotics makes this bacterium difficult to treat [2]. MTB mostly causes lung infections, but also infects other organs, such as the kidneys, lymph nodes, joints, vertebral column, and even the spleen.

Mycolic acids are hydroxyl fatty acids with long alkyl side chains. They are found in the cell wall of Mycolata taxon, which includes MTB, and increase resistance against chemical damage and dehydration, allowing the bacteria to readily grow inside macrophages and protect themselves against host immune systems. Mycolic acids are crucial for bacterial existence and maintenance of pathogenicity. MTB expresses and secretes three closely related mycolyl transferases with masses between 30 kDa and 35 kDa, known as antigen-85 protein complexes (Ag85A, Ag85B, and Ag85C), which are involved in the biogenesis of MTB cell walls. Given their vital role in cell wall synthesis, they are considered an ideal target for therapeutic treatment against TB [3]. These proteins catalyze the transfer of mycolic acid from one trehalose 6-monomycolate (TMM) to another, forming trehalose 6,6-dimycolate (DMM) and free trehalose.

Antimicrobial peptides (AMPs) are short stretches of amino acids made up of less than 50 amino acid residues with a molecular weight of between 2 kDa and 12 kDa and a broad spectrum of antimicrobial activity. AMPs can be cationic, anionic, or neutral are a key component of the innate immune system present in multicellular organisms [4]. A large number of AMP families are produced in animals, with the two main families called defensins and cathelicidins. AMPs exhibit broad spectrum antimicrobial activity. One advantage is that microbes do not easily gain resistance to them due to their nonspecific binding to the cell membrane. Therefore, they can be used to treat infections caused by organisms that are resistant to conventional antibiotics, i.e., drug resistant. Dermcidin is an AMP encoded by the DCD gene (NM_001300854.1) in humans. It is secreted by sweat gland cells to provide a barrier function [5]. Dermcidin display a broad spectrum of antimicrobial activity against both Gram-positive and Gram-negative bacterial pathogens [6]. The present study was undertaken in order to understand the inhibitory effect of dermcidin on mycolyl transferase by means of in silico studies.

  Materials and methods Top

System requirements

The entire study was performed on a Sony Vaio Laptop with a 2.67 GHz processor, 4 GB RAM, and a 320 GB hard drive and a Windows Professional operating system. All software used for the present study were open-source tools available free for download.

Receptor molecule fibronectin-binding protein A (FbpA)

MTB expresses three mycolyl transferase enzymes that are closely related and together constitute an Ag85 complex. They are involved in cell wall synthesis by catalyzing the reaction between TMM and mycolic acid to form DMM, also called a cord factor, which is a structure necessary for maintaining cell wall integrity [7]. They are also involved in transformation of mycolic acid from TMM to arabinogalactan for the peptidoglycan layer of the cell wall, ultimately forming an arabinogalactan-mycolate polymer [8]. The structure of mycolyl transferase, also known as FbpA, was retrieved from the Protein Data Bank (PDB ID: 1SFR) [9]. FbpA is an enzyme with a molecular weight of 35.68 kDa, a pI of 6.51, and consisting of 304 amino acid residues. Data were also obtained from the tuberculist database (http://tuberculist.epfl.ch/; RV number: 3804c) [9],[10].

Ligand molecule

In the present study, dermcidin AMP was used as a ligand molecule. Dermcidin is a peptide of 110 amino acid residues that is cleaved by proteases present in human sweat to produce peptides of varying length and diverse function. The dermcidin C-terminal peptide possesses antibacterial and antifungal activity and is constitutively expressed in sweat, while the N-terminal peptide promotes survival of neural cells under conditions of severe oxidative stress. The amino acid sequence of the C-terminal peptide is NH2–SSLLEKGLDGAKKAVGGLGKLGKDAVEDLESVGK–COOH and was used in the present study to generate the protein model. The protein model was generated using the SWISS MODEL online web tool. (http://swissmodel.expasy.org/) [11]. SWISS MODEL is a fully automated protein structure homology modeling server. Based upon query coverage and a Global Model Quality Estimation (GMQE) score, a template was selected to model the protein. GMQE estimates the quality of the generated protein model, which combines properties from the target-template alignment [12].

Docking software

Four different docking software tools were used. Discovery Studio 4.1 is software developed by Accelrys (Dassault Systemes, BIOVIA Corp., San Diego, CA, USA) for creating small-molecule structures, as well as macromolecules. A Molecular Graphics Laboratory (MGL) tool (The Scripps Research Institute, La Jolla, CA, USA) was used for the visualization and analysis of molecular structures. AutoDock (The Scripps Research Institute) is a docking tool used to predict the interaction between small molecules, such as drug compounds or substrates, with the three-dimensional structure of a molecule. Docking between Mycolyl transferase (FbpA-receptor) and dermcidin AMP (ligand) was performed using AutoDock Vina. Docked structures were visualized using the open source, molecular visualization system PyMol 1.7.4 (Schrödinger, LLC, Cambridge, MA, USA).

Docking protocol

The receptor and ligand molecules were prepared for docking using Discovery Studio 4.1 (Dassault Systèmes BIOVIA, Discovery Studio Modeling Environment, Release 4.5, 2015). FbpA molecule was opened using Discovery Studio, after which water molecules and other bound ligand molecules were removed. The binding site was defined after adding hydrogen molecules to the FbpA receptor molecule in order to describe the receptor cavity. FbpA (receptor) and dermcidin (ligand) molecules were saved as PDB files, then opened in the AutoDockTools portion of MGL 1.5.6 (The Scripps Research Institute) in order to prepare the receptor and ligand molecule for docking. Docking was performed using AutoDock Vina [13] and docked models visualized using PyMol (Schrödinger, LLC).

  Results and discussion Top

Model validation

The AMP selected in the present study is dermcidin, which was confirmed by matrix-assisted laser desorption/ionization mass spectrometry analysis. Dermcidin is an AMP with 110 amino acid residues. Following secretion, it undergoes postsecretory proteolytic processing by proteases, creating small peptides of between 25 and 48 amino acid residues [14]. A model of dermcidin was generated based on 100% query coverage using the template molecule (PDB ID: 1KSG) [15]. The generated model was validated based on parameters mentioned in Section “Materials and methods”.

Ramchandran plot

The Ramchandran plot for the template molecule was retrieved from the PDB and the plot for the modeled protein was also generated. Structural evaluation was performed using RAMPAGE (http://mordred.bioc.cam.ac.uk/~rapper/rampage.php). Ramchandran plot analysis for the modeled protein revealed that 98% of the residues localized within favored regions, suggesting good quality of the model ([Figure 1]).
Figure 1: Ramchandran plots for (A) 1KSG (template) and (B) modeled dermcidin molecule.

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GMQE score

The GMQE score is generated by the target-template alignment of the modeled protein and the value is expressed as a number between zero and one. Higher GMQE scores indicate increased structural reliability [11]. The GMQE score for the modeled protein was found to be 0.98, which indicates good model accuracy.

Structural visualization

Structures of the template molecule and modeled molecules were visualized as described in Section “Materials and methods” and compared for structural variation ([Figure 2]).
Figure 2: Structural comparison of models (A) 1KSG and (B) model.

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Enzyme active site

The Ag85A, Ag85B, and Ag85C complexes have mycolyl transferase activity that catalyze glycolipid synthesis of the mycobacterial cell wall, called the cord factor, and are also known as FbpA, FbpB, and FbpC. FbpA is a trimer consisting of three identical polypeptide chains (A, B, and C) of 304 amino acid residues each ([Figure 3]A). In a previous study by Ronning et al. (2004) [9], OSG-octylthioglucoside was used as a substrate for Ag85C, given its similarity to a portion of the AG85C natural substrate trehalose-monomycolate. They studied the structure of Ag85C using OSG-octylthioglucoside as a substrate in order to obtain information regarding substrate binding and its interaction with the active site. Data revealed two types of interactions taking place between OSG-octylthioglucoside and Ag85C: van der Waals contacts and hydrogen bonds. Trp-158, Leu-161, Ile-163, and Ala-165 are involved in van der Waals interactions between OSG-octylthioglucoside and Ag85C. Gly-39 and Arg-41 are involved in the formation of hydrogen bonds with the glucose moiety of the substrate and the side chains of Asp-38, Asn-52, and Trp-262 residues located within 5.4Å of the substrate, as well as interactions with the disaccharide ring of the carbohydrate moiety [9]. Here, FbpA was selected as a target for ligand binding. The FbpA receptor cavity is shown in [Figure 3]B. The amino acid residues involved in formation of the receptor cavity are shown in [Table 1]. Approximately 45 amino acid residues are involved in the formation of the FbpA receptor cavity. Over 50% of the residues have hydrophobic side chains and are involved in hydrophobic interactions. The receptor cavity is being formed between chains A and B ([Figure 3]B), so only the residues of chains A and B are involved in receptor cavity formation.
Figure 3: FbpA (A) mycolyl transferase (PDB ID: 1SFR) chains A, B, and C. (B) Formation of receptor cavity in mycolyl transferase

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Table 1: Amino acid residues involved in formation of the receptor cavity.

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Docking study

The docking of FbpA with dermcidin AMP was performed by AutoDock Vina, generating nine different poses ([Table 2]). The pose with best fit was selected based on having the lowest binding energy associated with interactions with the ligand molecule. In the present study, 5.6 kcal/mol was found to be the lowest energy value relative to other results and, therefore, was considered to represent the best fit model. The docked model was visualized using PyMol ([Figure 4]). FbpA binding to dermcidin involves 13 hydrogen bonds between substrate and FbpA residues ([Table 3]). Of these 13 residues, six are involved with receptor cavity formation, confirming localization of dermcidin binding within the receptor cavity ([Table 3]) in the cleft between FbpA chains A and B ([Figure 4]). FbpA docking with OSG-octylthioglucoside displayed nine different areas of interaction, with the lowest binding energy measured at 7.4kcal/mol. Residues involved in interactions between FbpA and OSG-octylthioglucoside were Leu-42, Ser-126, His-262, and Trp-264. Of these four residues, two residues, i.e. Leu-42 and Ser-126 are located in the receptor cavity. This confirms the interaction of OSG-octylthioglucoside molecule with the receptor cavity. Comparison of residues involved in receptor cavity formation and those involved in hydrogen bonding between FbpA and dermcidin or OSG-octylthioglucoside indicated that both ligands bind in the receptor cavity. Therefore, we can postulate that the binding of dermcidin in the receptor cavity might affect binding of its natural substrate, mycolic acid, and, therefore, interfere in MTB cell wall synthesis.
Figure 4: Visualization of docking between (A) FbpA and OSG-octylthioglucoside and (B) FbpA and the modeled dermcidin molecule.

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Table 2: Nine poses with energy calculations and RMSD values.

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Table 3: Residue interactions between receptor and ligand.

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  Conclusion Top

The present study concludes that dermcidin binds with FbpA to inhibit mycolic acid transfer to the peptidoglycan layer of the cell wall, interfering with cell wall synthesis.

Conflicts of interest

The authors declare no conflicts of interest.


The authors are thankful to the Indian Council of Medical Research, New Delhi, India (IRIS ID: 2011-13260) for providing financial support and the Department of Integrated Biotechnology, Ashok and Rita Patel Institute of Integrated Study and Research in Biotechnology, Allied Sciences, and the Charutar Vidya Mandal institute. We also acknowledge Dr. Sunil Trivedi, HOD, Microbiology, Shree Krishna Medical Hospital, Anand Agricultural University and SICART, India, for providing technical support.

  References Top

Global Tuberculosis Control 2014, WHO Geneva (2014), <www.who.int/tb/publications/global_report> [accessed 16.07.2015].  Back to cited text no. 1
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H.G. Wiker, M. Harboe, The antigen 85 complex: a major secretion product of Mycobacterium tuberculosis, Microbiol.Rev. 56 (1992) 648–666.  Back to cited text no. 3
T. Ganz, Defensins: antimicrobial peptides of innate immunity, Nat. Rev. Immunol. 3 (2003) 710–720.  Back to cited text no. 4
B. Schittek, R. Hipfel, B. Sauer, et al, Dermcidin: a novel human antibiotic peptide secreted by sweat glands, Nat. Immunol. 2 (2001) 1133–1137.  Back to cited text no. 5
H. Steffen, S. Rieg, I. Wiedemann, et al, Naturally processed dermcidin-derived peptides do not permeabilize bacterial membranes and kill microorganisms irrespective of their charge, Antimicrob. Agents Chemother. 50 (2006) 2608–2620.  Back to cited text no. 6
J.T. Belisle, V.D. Vissa, T. Sievert, et al, Role of the major antigen of Mycobacterium tuberculosis in cell wall biogenesis, Science 276 (1997) 1420–1422.  Back to cited text no. 7
K. Takayama, C. Wang, G.S. Besra, Pathway to synthesis and processing of mycolic acids in Mycobacterium tuberculosis, Clin. Microbiol. Rev. 18 (2005) 81–101.  Back to cited text no. 8
D.R. Ronning, V. Vissa, G.S. Besra, et al, Mycobacterium tuberculosis antigen 85A and 85C structures confirm binding orientation and conserved substrate specificity, J. Biol. Chem.279 (2004) 36771–39777.  Back to cited text no. 9
L. Kremer, W.N. Maughan, R.A. Wilson, et al, The M. tuberculosis antigen 85 complex and mycolyltransferase activity, Lett. Appl. Microbiol. 34 (2002) 33–237.  Back to cited text no. 10
B. Macro, B. Stefan, W. Andrew, et al, Swiss-model: modelling protein tertiary and quaternary structure using evolutionary information, Nucl. Acids Res. 42 (2014) 252–258.  Back to cited text no. 11
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S. Rieg, S. Seeber, H. Steffen, et al, Generation of multiple stable dermcidin-derived antimicrobial peptides in sweat of different body sites,J. Invest. Dermatol. 126 (2006) 354–365.  Back to cited text no. 14
H.B. Michael, R. Louis, P. Roversi, A. Wittinghofer, R.C. Hillig, The complex of Arl-GTP and PDEd: From structure to function, EMBO J. 21 (2002) 2098–2106.  Back to cited text no. 15


  [Figure 1], [Figure 2], [Figure 3], [Figure 4]

  [Table 1], [Table 2], [Table 3]

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