Oral Presentation 2014 International Biophysics Congress

Molecular tools for targeted covalent labeling and light-induced activation of biopolymers (#16)

Saulius Klimasauskas 1 , Grazvydas Lukinavicius 1 , Rasa Rakauskaite 1 , Viktoras Masevicius 1 , Edita Kriukiene 1 , Giedre Urbanaviciute 1 , Audrone Ruksenaite 1
  1. Institute of Biotechnology, Vilnius University, Vilnius, Lithuania

Functional analysis of biological systems requires the ability to visualize and manipulate selected biomolecules in their complex environments, which is often a challenging task.  Here we present two novel platforms for targeted covalent manipulation of biopolymers. The first approach relies on methyltransferase enzymes, which catalyze highly specific methyl group transfers from the ubiquitous cofactor S-adenosylmethionine (AdoMet) to a multitude of biological targets in the cell. We redesigned the methyltransferase reaction for covalent transfer of larger chemical entities by engineering the catalytic center and employing synthetic AdoMet analogs with transferable reporter or functional groups1,2,3. The new approach provides a new enabling tool for a highly targeted functionalization and labeling of natural DNA and tRNA, miRNA, rRNA4,5,6,7. We have also developed a universal technique for incorporation of a genetically-encoded photocaged selenocysteine residue into any position of a recombinant protein. Selenocysteine is of significant technological importance as a component of both natural proteins and designed biocatalysts, however the availability of such proteins is hampered by technical limitations. We demonstrate that photo-decaging of the incorporated residue unmasks the unique selenol functionality, which may serve as an efficient site for light-induced protein dimerization inside yeast cells or permit site-specific labeling of the recombinant protein 8. Altogether, the proposed approaches offer an unprecedented flexibility in chemical manipulation of biomolecules for cellular imaging, epigenome profiling, single-molecule genomics4,7 and spatiotemporal control of protein function 8.

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  2. Lukinavičius et al. (2007) J. Am. Chem. Soc. 129: 2758–2759.
  3. Klimašauskas S. & Weinhold E. (2007) Trends Biotechnol. 25: 99–104.
  4. Neely et at., (2010) Chem. Sci., 2010, 1: 453-460.
  5. Tomkuvienė, et al. (2012) Nucleic Acids Res., 40: 6765-6773.
  6. Lukinavičius et al. (2012) Nucleic Acids Res., 40: 11594–11602.
  7. Kriukienė, et al. (2013) Nat. Commun., 4: 2190.
  8. Rakauskaitė et al., in preparation.