Mycobacterial peptide deformylase

Abstract

The present invention relates to the design of the Antisense-oligonucleotide complementary to the specific region of peptide deformylase gene from Mycobacterium tuberculosis . The use of this Antisense-oligonucleotide on mycobacterial culture inhibits the production of the peptide deformylase enzyme by hybridizing within the region, which is found to be responsible for maintaining stability as well as retaining the functionality of the enzyme and thus in turn affecting the growth of the cells. This invention also establishes the essentiality of the peptide deformylase enzyme in mycobacteria and claims it as a drug target in this microorganism.

1 . A purified polynucleotide sequence comprising an insertion sequence of the mycobacterial peptide deformylase [def] gene represented by the nucleotide of sequence ID No. 14. 
     
     
         2 . A purified polynucleotide sequence as claimed in  claim 1 , wherein said sequence is useful as a potential drug target against mycobacteria. 
     
     
         3 . A polypeptide encoded by the polynucleotide sequence of  claim 1 , wherein said polypeptide comprises 12 amino acids represented by XTXRRRGVVINP, wherein X=any one of the 20 known amino acids. 
     
     
         4 . A polypeptide as claimed in  claim 3 , wherein the sequence is 90 to 95% similar to the insertion amino acid sequence of the mycobacterial peptide deformylase [def] gene in  M. tuberculosis, M. smegmatis, M. bovis, M. avium  and  M. leprae.    
     
     
         5 . An antisense oligonucleotide complementary to the nucleotide sequence represented by sequence ID No. 14. 
     
     
         6 . An oligonucleotide as claimed in  claim 5 , wherein said oligonucleotide is characterized in that said oligonucleotide consists of one or more phosphorothioate modified oligodeoxynucleotide. 
     
     
         7 . An oligonucleotide as claimed in  claim 5 , wherein said oligonucleotide inhibits the production of the enzyme peptide deformylase by hybridizing within the short region of mycobacterial peptide deformylase (def) gene. 
     
     
         8 . An oligonucleotide as claimed in  claim 5 , wherein said oligonucleotide is a potential drug against mycobacteria 
     
     
         9 . A process for the preparation of an antisense oligonucleotide as claimed in  claim 5 , the said process comprising the steps of:
 a. Isolating polynucleotide sequence from  M. tuberculosis  comprising nucleic acid sequence (594 bp) encoding a polypeptide (197 amino acids) having peptide deformylase activity wherein, the polypeptide is present in different mycobacterial species like  M. tuberculosis, M. smegmatis, M. bovis, M. avium, M. leprae  represented by SEQ ID NO: 8, 9, 10, 11, 12 and having at least 90 to 95%, sequence similarity among themselves;   b. Identifying a region within mycobacterial peptide deformylase enzyme isolated from step (a) represented by polynucleotide sequence ID No. 14 and amino acid sequences 74 to 85 involved in maintaining the enzymatic stability and functionality, the said region being conserved in all of the mycobacterial species as given in step (a);   c. preparing an antisense oligonucleotide or its permissive modifications, against the conserved region of peptide deformylase enzyme, as identified in step (b);   d. Inhibiting the enzyme activity as well as growth of the mycobacteria using the antisense oligonucleotide prepared in step (c).   
     
     
         10 . The polynucleotide insertion sequence as claimed in  claim 1 , useful as a potential drug target against mycobacteria. 
     
     
         11 . The polypedide encoded by the def gene of mycobacteria as claimed in  claim 3 , useful as a potential drug target against mycobacteria. 
     
     
         12 . The oligonucleotide as claimed in  claim 5 , useful for inhibiting the activity and growth of mycobacteria. 
     
     
         13 . A pharmaceutical composition useful for the treatment of tuberculosis comprising an antisense oligonucleotide or any biotic/abiotic compound(s) that interacts with the insertion sequence of the mycobacterial peptide deformylase [def] gene of  claim 1 , optionally along with pharmaceutically acceptable carriers, additives or diluents.