Oral Presentation 10th Australian Peptide Conference 2013

A strategy for stabilizing weak interactions of posttranslationally modified proteins (#69)

Nam K. Chu 1 , Konstantinos Tripsianes 2 , Anders Friberg 2 , Michael Sattler 2 , Christian F.W. Becker 1
  1. University of Vienna, Vienna, Austria
  2. Technische Universität München, Chemistry, Munich, Germany

Survival of motor neuron (SMN) protein is part of the SMN complex that mediates assembly of ribonucleoprotein complexes1. SMN protein contains a central Tudor domain that recognizes symmetrically dimethylated arginine residues (sDMA), leading to transient interactions with Sm proteins. Due to the weak but specific interaction of SMN Tudor with sDMA (KD ~ 0.5 mM) a large excess of free sDMA had to be used in previous structural studies of the SMN Tudor-sDMA complex2. Such conditions are not ideal for structure analyses of weak interactions and covalent coupling of the two interacting moieties can shift the equilibrium to the bound state by lowering the entropic cost of binding.

Here sDMA is covalently linked to SMN Tudor via expressed protein ligation, using a flexible glycine linker3. NMR analyses and ligation yields allow assessment of the ideal linker length and enable structural analysis of a weak interaction at moderate concentrations. The described strategy can help to overcome current challenges in structural analysis of such weak, transient interactions often mediated by posttranslational modifications.

  1. Pellizzoni, L.; Yong, J.; Dreyfuss, G. Science 2002, 298, 1775-9.
  2. Tripsianes, K.; Madl, T.; Machyna, M.; Fessas, D.; Englbrecht, C.; Fischer, U.; Neugebauer, K. M.; Sattler, M. Nat. Struct. Mol. Biol. 2011, 18, 1414-20.
  3. Muir, T. W.; Sondhi, D.; Cole, P. A. Proc. Natl. Acad. Sci. U.S.A. 1998, 95, 6705-10.