Inteins and protein splicing

summary:   Protein splicing is a post-translational process that results in excision of an internal protein region (intein) and ligation of its flanking sequences (exteins). Protein splicing does not require cofactors of host's enzymes: it is catalyzed by itself through the internal domain (Hint-domain) [a]. It can be explained by tertiary structure of intein. The distance between the N and C ends of the intein is comparable with the length of a covalent bond (about 1.4 Å). Hint-domain plays an important role in autocatalytic cleavage of the intein and subsequent ligation of the exteins. Interestingly, that the Hint-domain catalytic center structurally is similar to the active center of serine protease.

The main aims of our work was to create intein-based chimeric protein for quick purification of the human growth hormone (HGH). I made a chimeric protein consisting of a short N-terminal peptide, the Mxe GyrA intein, and human growth hormone between them. I did quick affinity purification by the intein, and then induce self-cleavage of the intein, that releases human growth hormone in a pure buffer [bc].

LOCATION: Institute of molecular biology and genetics NAS of Ukraine, Kiev

      Obtained degree and awards:

     Publications:      

  1. Starokadomskyy P.L., Okunev O.V. ,  Irodov D.M., Kordium V.A. Utilization of Protein Splicing for Purification of the Human Growth Hormone // Molecular biology (Moscow), 2008, Vol. 42, No. 6, pp. 966-972. PMID: 19140330
  2. Starokadomskyy P.L., Okunev O.V., Dubey L.V. Utilizing of protein splicing phenomenon for optimization of obtaining and purification of the recombinant human growth hormone  // Eur J Med Res., 2006, Oct. 8, vol.11 (17th  European students Conference, 8-12 October 2006, Berlin) p. 128-129. (link)
  3. Starokadomskyy P.L. Protein splicing // Molecular biology (Moscow), 2007, vol. 41, N2, p. 278-293. PMID: 17514899
  4. Starokadomskyy P.L., Dubey I.Ya., Okunev O.V., Irodov D.M. Construction of a Chimeric Intein-Containing Protein and the Search for Conditions for Its Cleavage // Cytology and Genetics, 2007, vol. 41, N2, p. 3-11 (PMID: 17494337)
  5. Starokadomskii P.L. Protein splicing // The Ukrainian biochemical journal, 2005, №4, p.14-29.  (PMID: 16568601)

 

   SOME DATA:

   1. Scheme of protein cis-splicing            

  2. Scheme of protein trans-splicing

  3. Comparative analyses of the active centers of serine protease and Mxe GyrA intein

 



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