Technical Highlight - November 2010
Short description: Complexes are very difficult to solve in solution but are important biologically. Combining multiple approaches is the way forward.
Dimeric and multiprotein complexes are difficult to tackle by NMR, yet it is an important problem to solve because such a high proportion of proteins are thought to be in complex in the cell. For homodimers, it is a challenge to distinguish intramolecular NOEs from intermolecular ones. But incorporating additional information from the latest NMR techniques has brought routine solution of dimeric structures within reach.
One successful approach used by a couple of groups, including Michael Kennedy and colleagues from PSI NESG consortium, is to combine electron paramagnetic resonance (EPR)–based double electron-electron resonance (DEER) measurements with paramagnetic resonance enhancements (PRE) and NOE information. EPR-based DEER and PRE measurements can provide additional distance constraints over a range of 10–70 Å (ref. 1).
Kennedy's group used such mid- to long-range data to solve the structure of the homodimeric spore-coat assembly protein Dsy0195 from Desulfitobacterium hafniense. By introducing spin labels through mutagenesis, they were able to use PRE and DEER along with conventional NMR techniques to solve the structure using the automatic NMR structure determination program CYANA.
James Prestegard's team, also from PSI NESG, have applied an alternative approach to resolving homodimeric structures, using residual dipolar coupling (RDC) information 2 . They tackled SeR13, a protein from Staphylococcus epidermidis, which is a weakly associated homdimer. Their tactic was to solve the monomeric structure first, then use RDC constraints to dock the dimer. They checked the resulting structure using paramagnetic surface and chemical shift perturbations.
Both approaches show the power of combining additional data with standard NMR techniques and look likely to usher in an era of multiprotein solution structures.
Y. Yang, T. A. Ramelot, R. M. McCarrick, S. Ni, E. A. Feldmann et al. Combining NMR and EPR methods for homodimer protein structure determination.
J. Am. Chem. Soc. 132, 11910-11913 (2010). doi:10.1021/ja105080h
H.-W. Lee, G. Wylie, S. Bansal, X. Wang, A. W. Barb et al. Three-dimensional structure of the weakly associated protein homodimer SeR13 using RDCs and paramagnetic surface mapping.
Prot. Sci. 19, 1673-1685 (2010). doi:10.1002/pro.447