International Journal of Biochemistry and Biophysics 3(2): 13-23, 2015 http://www.hrpub.org DOI: 10.13189/ijbb.2015.030202 Insight into the LGP2 Helicase Domain – An in Silico Approach Subhomoi Borkotoky, Chetan Kumar Meena, Aswathy Soman, Ayaluru Murali * Bioinformatics Centre, School of Life Sciences, Pondicherry University, Puducherry, India Copyright © 2015 Horizon Research Publishing All rights reserved. Abstract LGP2, a member of retinoic acid inducible gene–I like receptors (RLR), encoded by the gene DHX58 in human induces antiviral response against many RNA viruses. The LGP2 shares a considerable similarity to the amino acid sequence of Hef Helicase Domain, the helicase domain of RIG-I-like protein helicase-associated endonuclease (Hef), which is involved in RNA binding. Earlier studies suggest that RLR contains a conserved C-terminal domain (CTD), which is responsible for the binding specificity to the viral RNAs and C-terminal domain of LGP2 also takes part in RNA binding. The present study is aimed to explore the interactions of LGP2 and RNA, thereby finding the crucial residues for the interaction, with the help of in-silico tools. The predicted crucial residues were validated by docking and molecular dynamics simulation studies as well as by fitting the model on a LGP2 density map. Our results in this study suggest that the residues in the helicase domain of LGP2 are crucial for RNA binding and it is positioned around the groove region of LGP2. Though earlier experimental studies identified the RNA binding residues, but our in silico binding studies with the full length LGP2 predicted some additional residues that can be valuable Keywords LGP2, Homology Modeling, Molecular Docking, Molecular Dynamics Simulation, Flexible Fitting 1. Introduction The first line of defense, innate immune response is dependent on pattern recognition receptors; one class of pattern recognition receptors is toll like receptors, found on the cell surface or endosomes in effector cells, that recognize pathogen associated molecular pattern typically located on the surface of pathogen cells [1], whereas RIG-I like receptors (RLRs) are in the cytoplasm. The RLR family, within superfamily 2 (SF2) RNA helicases, include RIG-I (retinoic acid-inducible gene-I), MDA5 (melanoma-differentiation-associated gene 5), and LGP2 (Laboratory of Genetics and Physiology 2) and are closely related to DEAD-box helicases that can sense RNAs to initiate antiviral responses [2, 3]. In human, LGP2 is encoded by DHX58 gene [4]. Though LGP2 is quite similar to other RLR family members RIG-I and MDA-5, it lacks the signature N-terminal caspase activation and recruitment domain (CARD) (Fig. 1) that is required for signaling [5]. Different studies defined LGP2 as a positive as well as negative regulator of RIG-I and MDA5-mediated viral recognition [6, 7]. Further, it was demonstrated that LGP2 protein produced in insect cells can bind both single- and double-stranded RNA (ss- and ds-RNA), with higher affinity for dsRNA and significant change in the conformation of the LGP2 monomer to a stable dimer takes place when it was complexed to a dsRNA [8]. The RLR contains a conserved C-terminal domain (CTD), which is responsible for the binding specificity to the viral RNAs. The solution structure of LGP2 (PDB ID: 2RQA) C-terminal domain (CTD) (546 – 678) has shown that it binds to dsRNA and 5ʹppp-ssRNA with higher affinity than CTD of RIG-I [9]. A similar view was also reported in the 2.0-Å resolution crystal structure of human LGP2 CTD and 8-bp dsRNA complex (PDB ID: 3EQT), which was also supported by gel filtration chromatography and analytical ultracentrifugation studies [10]. The helicase domain of RIG-I-like protein, helicase-associated endonuclease (Hef), consists of three structural sub-domains that share common conserved sequences like other SF2 helicase members, e.g. RIG-I and DEAD-box. The Hef helicase domain (Hhd, residues 1–494) contains seven helicase motifs that were spread in sub-domains 1 and 3 [11, 12]. The crystal structure of Hhd resembles a partially opened clamp with a concave grove associated with its lower portion [11]. Murali et al [8] , using electron microscopy and single particle analysis reported the low resolution structure of LGP2. By superimposing the crystal structure of Hef helicase domain into the electron density map of LGP2, it was observed that LGP2 shares a considerable structural similarity with the three domains of Hhd [7]. Further LGP2 also shares a considerable sequence similarity with Hhd (Figs. 1 and 3.i). It was also reported that the RNA binding motifs that were observed in Hhd [11] were also conserved in LGP2 (Fig. 3.i) and that they aligned in the groove part of the LGP2 structure when the crystal structure of Hhd is superimposed on the LGP2 density map [8].