Analysis and modeling of heat-labile enterotoxins of Escherichia coli suggests a novel space with insights into receptor preference M. Krishna Raja, Asit Ranjan Ghosh, S. Vino and S. Sajitha Lulu* School of Bio Sciences and Technology, VIT University, Vellore, India Communicated by Ramaswamy H. Sarma (Received 20 January 2014; accepted 4 October 2014) Features of heat-labile enterotoxins of Escherichia coli which make them fit to use as novel receptors for antidiarrheals are not completely explored. Data-set of 14 different serovars of enterotoxigenic Escherichia coli producing heat-labile toxins were taken from NCBI Genbank database and used in the study. Sequence analysis showed mutations in different subunits and also at their interface residues. As these toxins lack crystallography structures, homology modeling using Modeller 9.11 led to the structural approximation for the E. coli producing heat-labile toxins. Interaction of modeled toxin subunits with proanthocyanidin, an antidiarrheal showed several strong hydrogen bonding interactions at the cost of minimized energy. The hits were subsequently characterized by molecular dynamics simulation studies to monitor their binding stabilities. This study looks into novel space where the ligand can choose the receptor preference not as a whole but as an individual subunit. Mutation at interface residues and interaction among subunits along with the binding of ligand to individual subunits would help to design a non-toxic labile toxin and also to improve the therapeutics. Keywords: receptor; labile toxin; interface residue; mutation 1. Introduction Enterotoxigenic Escherichia coli (ETEC) play a critical role in causing diarrheal-related illness among mam- mals (Levine, 1987). It causes travelers’ diarrhea mainly in the developing world (Wang et al., 2010) and also causes diarrhea in infants, children, and adults leading to global health problem. It emphasizes the need to understand and analyze the role of ETEC toxin genes in humans which has a heterohexamer (AB5) complex consisting of catalytically active monomeric A subunit (LTA) and pentameric B subunits (LTB) (Merritt & Hol, 1995). Heat-labile (LT) enterotoxin is one of the essential components in disease pathogenesis resulting in gut lumen dehydration (Nataro & Kaper, 1998). In addition to pathogenicity expression of LT, colonization is an added advantage (Allen, Randolph, & Fleckenstein, 2006; Berberov et al., 2004). Vaccine development is facing hurdles due to the variability of antigens among strains (Svennerholm & Tobias, 2008). It is well documented that the mutation has an impact on toxicity, adjuvant activity, cytotonic effect, cAMP levels, and ADP ribosylation (Rodrigues et al., 2011). Significance of mutation can be deciphered from reports like mutation in cysteine residues which results in degradation of LTA by proteases and thereby reduc- tion in cAMP production (Okamoto, Nomura, Fujii, & Yamanaka, 1998). Therefore, the present study enumerates the necessity of analyzing the subunit sequence for mutation, interface residues, and interaction of ligands with individual sub- units which would be useful for developing drugs for treatment and prevention. 2. Methods 2.1. LT data-set Complete sequences of heat-labile enterotoxin (LT) of E. coli were taken from the Genbank database. Among the several hits found, the complete sequences were selected from both the A and B subunit toxin. As illus- trated in Figure 1, using keyword search in Genbank database, the respective hits 14 for LTA and 71 for LTB were obtained as search results (Table 1). For LTA, the 14 hits representing E. coli were selected, while for LTB the first 14 hits for full length sequence were selected for the data-set. 2.2. Multiple sequence alignment of LTA and LTB Multiple sequence alignment (MSA) was performed using clustalW2 protein with the substitution matrix PAM, extension penalty of .20, and gap-opening penalty of 10. 1LTS available in the Brookhaven protein data bank (PDB) database (Abola, Bernstein, & Koetzle, *Corresponding author. Email: ssajithalulu@vit.ac.in © 2014 Taylor & Francis Journal of Biomolecular Structure and Dynamics, 2014 http://dx.doi.org/10.1080/07391102.2014.974073