SITE DIRECTED MUTAGENESIS OF HUMAN INTERLEUKIN-2 GENE TO INCREASE THE STABILITY OF THE GENE PRODUCT- A BIOINFORMATICS APPROACH

Ashwin Kumar Dakshinamurthi¹, Manthira Vasagam Chidambaram², Vivek Anand Manivel³, Swaminathan Detchanamurthy⁴*
¹Department of Biotechnology, Sri Venkateswara College of Engineering, Sriperumbudur, Tamilnadu, India
²Department of Biotechnology, Sri Venkateswara College of Engineering, Sriperumbudur, Tamilnadu, India
³Department of Biotechnology, Sri Venkateswara College of Engineering, Sriperumbudur, Tamilnadu, India
⁴Department of Chemical and Process Engineering, University of Canterbury, Private Bag: 4800, Christchurch, New Zealand
* Corresponding Author : swami_biochem@yahoo.co.uk

Received : -     Accepted : -     Published : 21-12-2009
Volume : 1     Issue : 2       Pages : 4 - 13
Int J Bioinformatics Res 1.2 (2009):4-13
DOI : http://dx.doi.org/10.9735/0975-3087.1.2.4-13

Keywords : IL-2, in-silico, site directed mutagenesis, cytokine, bioinformatics
Conflict of Interest : None declared

Interleukin-2 (IL-2) is an immunoregulatory cytokine whose biological effects are mediated through interaction with specific receptors on the surface of target cells. Due to its presumed role in generating a normal immune response, IL-2 is being evaluated for the treatment of a variety of tumors, in addition to infectious diseases. Main drawback of human IL-2 is that the molecule is relatively unstable. Therefore, with the objective of increasing the stability of the molecule, site directed mutagenesis of human IL-2 gene was carried out. Early studies indicated that mutations at three Cysteine residues (58, 105, 125) which are in the active sites of human IL-2 resulted in the reduced stability as well as the biological activity of the molecule. Therefore, mutations were carried out at the positions of amino acid other than the receptor binding sites at 111Valine to Arginine, 117Lysine to Glutamine and 133 Threonine to Asparagine of the human sequence by comparing it with the bovine sequence which has higher stability than the human counterpart, using SWISS PDB tool. To understand the biological activity of the mutated IL-2, energy minimization studies were carried out using SWISS-PDB. Docking studies were performed to check the reliability of the results using HEX DOCK, ARGUS LAB and PATCH DOCK between the IL-2 receptor and its mutated Ligand. These docking results also confirmed that the reliability of these mutated IL-2 gene. Stability, half life and ADME characteristics of these mutants can be studied in a detailed manner in the in vivo studies.

[1] Anderson D., Baker P.E., Cantrell M.A., Cerretti D.P., Cosman D., Gillis S., Larsen A., McKereghan K. (1986) Proceedings of the National Academy of Sciences of the United States of America, 83(10), 3223-3227.  
» CrossRef   » Google Scholar   » PubMed   » DOAJ   » CAS   » Scopus  

[2] Barr P.J., Buck C.D., Magnuson J.A., Magnuson N.S., Nissen M.S., Reevess R., Spies A.G and Weinberg A.D. (1986) PNAS, 83:10, 3228–3232.  
» CrossRef   » Google Scholar   » PubMed   » DOAJ   » CAS   » Scopus  

[3] Bartlett J.A., Blankenship K., Waskin H., Sebastian M., Austin A. and Weinhold K. (1991) Int Conf AIDS, 21(7), 220-225.  
» CrossRef   » Google Scholar   » PubMed   » DOAJ   » CAS   » Scopus  

[4] Berndt W.G., Chang D.Z., Smith K.A., Ciardelli T.L., (1994) Biochemistry, 33(21), 6571- 6577.  
» CrossRef   » Google Scholar   » PubMed   » DOAJ   » CAS   » Scopus  

[5] Berman H.M. (2000) Nucleic Acids Res. 28(1), 235–242.  
» CrossRef   » Google Scholar   » PubMed   » DOAJ   » CAS   » Scopus  

[6] Blattman J.N., Grayson J.M., Wherry E.J., Kaech S.M., Smith K.A. and Ahmed R. (2003) Nat Med, 9(5):540-547.  
» CrossRef   » Google Scholar   » PubMed   » DOAJ   » CAS   » Scopus  

[7] Lu S.D., Mark D.F. and Wang A. (1984) Science Journal, 22(4), 1431 – 1433.  
» CrossRef   » Google Scholar   » PubMed   » DOAJ   » CAS   » Scopus