Sumasridhar, Rudrammaji L. M. S , Balaji A , Chouhan T. R. S. Interaction of Ribosome Inactivating Proteins with Growth Factor Receptors - a Cheminformatic Approach. Biomed. Pharmacol. J.2008;1(1)

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Manuscript received on :April 12, 2008
Manuscript accepted on : June 04, 2008
Published online on: --

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Sumasridhar^1,2, L. M. S. Rudrammaji^³, Anna Balaji⁴ and T. R. S. Chouhan⁵

^¹Dr. MGR Educational and Research Institute (Deemed University), Chennai .

^²Department of Biochemistry, Dayananda Sagar College of Dental Sciences, Kumaraswamy layout, Bangalore .

^³Department of Biotechnology, Sri Jayachamarajendra College of Engineering, Mysore .

⁴Dayananda Sagar College of Pharmacy, Kumaraswamy layout, Bangalore .

⁵Director (Technical), Ethicamatrix, Hyderabad

Abstract

Protein interactions are the basis of life processes at the nano level. Most of the protein interactions are with other proteins. Thus efforts to recreate the network of protein-protein interactions are important for interpretation. Our study has such protein-protein interactions with a computational approach directly targeting ligand-receptor interactions through docking method and simulating the binding. Insilico studies on interaction of Ribosome Inactivating Proteins (RIP) with various growth factor receptors suggests “First approximation” for invitro studies. Docking of Ricin and Abrin polypeptide chains with Epidermal growth factor receptor and Fibroblast growth receptor was done by using Hex programme. This high throughput docking studies indicate results in terms of energy values. The interaction between the ligand and the receptor with minimal energy value is the best pair. Accordingly, the best docking results were obtained for interaction of the pairs,Ricin B chain - Epidermal growth factor receptor, Ricin B chain - Fibroblast growth factor receptor, Abrin - Epidermal growth factor receptor, Abrin - Fibroblast growth factor receptor with an Etotal values of -562.43, -427.77, -511.13 and -490.48 respectively.

Keywords

Epidermal Growth Factor Receptor (EGFR); Fibroblast Growth Factor Receptor (FGFR); Docking

Introduction

Many plants contain proteins that are capable of inactivating eukaryotic ribosomes, these are termed as “Ribosome-Inactivating Proteins” (RIPs)¹. Ricin from Ricinus communis and Abrin from Abrus precatorius belong to type II RIPs, both have got molecular similarity and similar mechanism of action ^2,3. They are composed of  2 chains A and B. The B chain helps in binding of proteins to cell surface receptors which are terminated with galactose containing oligosaccharides. The A chain is taken up into the cell and has N-glycosidase activity bringing about depurination of a specific adenine residue from the GAGA loop of 28S rRNA from 60S ribosome, thereby halting protein synthesis and leading to cell death.

Type-2 RIPs have a possible implication in therapeutics since these are more toxic to tumor cells than normal cells⁴. Hence these molecules were chosen for cheminformatic studies to understand protein-protein interactions, which offer a theoretical opportunity to develop antitumor drugs.

Cheminformatic  study is based on  computer guided system which is  capable of varying parameters. Modern drug designing efforts are exploiting at least three core technologies aimed at increasing the efficiency of finding drug leads viz., Genomics, High throughput Screening and Combinatorial Chemistry. Computational modeling has been developed and widely applied in better understanding of protein-protein interactions^5,6 . Computer simulations are done by studying macromolecular dynamics and free energy calculation methods⁷. Detailed energetics and structural knowledge of interactions between biomolecules is fundamental to understand the complex interactive mechanism that takes place in living organism and also to design drugs for blocking or modifying these interactions using molecular docking^8,9. In our study, we have used Rapid prototyping computational simulations using software tool Hex4.2. Using molecular dynamics and free energy calculations, the docking of ricin,and abrin was done with EGFR and FGFR. Growth factor receptors were selected for this insilico study because, in cancer, aberrant signaling can cause constitutive activation of growth factor receptors which may in turn influence key steps in the process of tumor invasion and metastasis. Involvement of EGFR and FGFR in tumor spread has indicated them as potential target receptors for antimetastatic studies.

Experimental

For the present study, Hex4.2 program was used which is an interactive molecular graphics program used to calculate the feasible docking pairs such as protein-protein (of our choice).  In Hex docking calculations, 3D parametric functions were used to encode both surface shape and electrostatic charge and their potential distribution.  Each property was represented by a vector of coefficients.  From Hex’s surface skin  model of protein  topology,  an  expression  for  a  docking  score  as  a function  of  six  degrees  of  freedom  in  a  rigid  body  docking  search with  suitable  scaling  factors was derived.  This docking score was interpreted as interaction energy.  This is a spherical polar approach where  the  molecules  were  relatively  rotated  to  generate  and  evaluate  good  docking  orientations  which  were effectively  a  six  dimensional   Fourier  Correlations.

Hex program was started by downloading and displaying the PDB structure of Ricin, Abrin and Epidermal growth factor receptor and fibroblast growth factor receptor. Docking was separately done with each RIP with EGFR and FGFR respectively. This procedure was repeated three times to ensure reproducible results.

Figure 1: Docking of Ricin with EGFR and FGFR Click here to View figure

Figure 2: Docking of Abrin with EGFR and FGFR Click here to View figure

There  were  four  stages  or  controls  in  the  program. The first control was the ‘orientation’.  Here  the  two molecules  were  oriented  with  their  favorable  positions  for  docking.  This was  followed  by  the  second  control,  ‘clustering’ where  the  program  uses  a  simple  clustering  algorithm  to  group  spatially  similar  docking  orientations.  The  third  control  of  this  program  was  ‘matching’ in which there  was  superposition  of  pair  of  proteins  ie,  RIP  and  the  receptor  and  a  search  for  maximum  similarity.  The  final  stage  was  ‘docking’  which  was  much  similar  to  matching. At this stage, the  surface  skin  coefficients  followed by docking  correlation  scores  at  each  of  the  specified  angular  and  intermolecular  increment were calculated.

Results and Discussions

The interaction studies reveal that the best docking results were for interaction of the pairs, Ricin B chain – Epidermal growth factor receptor,  Ricin B chain – Fibroblast growth factor receptor, Abrin – Epidermal growth factor receptor, Abrin – Fibroblast growth factor receptor with an E_total values of  -562.43  -427.77, -511.13 and -490.48 respectively as shown in the Table1.

Table 1: Docking Results

A large number of protein structures have been deposited into PDB, however only a small fraction of protein-protein complexes has been experimentally characterized so far. In this context theoretical prediction of protein-protein interactions is becoming critically important in structural biology. Using Hex4.2, we have used the 2 different RIPs as lead compounds which are prototype compounds having biological and pharmacological activities.  It is evident that the best interactive pattern is shown by ricin with EGFR with a very minimal energy value. The bumps of all the 4 interactions are –1 indicating that the stearic clashes are very minimal and thereby  better interaction. All these results  indicate that both the growth factor receptor can be targeted by these lead compounds in treatment of cancer.

Conclusion

The computational docking techniques are promising to be an essential tool in predicting the interactions of ligand-receptor complexes since it is cumbersome and very laborious to obtain crystal structures of protein complexes.  Ricin and Abrin can be powerful lead molecules which interact strongly with Epidermal and Fibroblast growth factor receptors as evidenced by the docking results.  The newly emerging field of cheminformatics with variety of software tools is addressing the needs of a broad community of scientists in First Approximation studies before venturing into invitro and invivo studies. Subsequently, these insilico data can be further extended into invitro studies and drug designing by targeting these lead molecules to the specific receptors which may have a positive implication in cancer therapy.

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