Abstract
Ras proteins are proto-oncogenes that function as molecular switches linking extracellular stimuli with an overlapping but distinctive range of biological outcomes. Although modulatable interactions between the membrane and the Ras C-terminal hypervariable region (HVR) harbouring the membrane anchor motifs enable signalling specificity to be determined by their location, it is becoming clear that the spatial orientation of different Ras proteins is also crucial for their functions. To reveal the orientation of the G-domain at membranes, we conducted an extensive study on different Ras isoforms anchored to model raft membranes. The results show that the G-domain mediates the Ras–membrane interaction by inducing different sets of preferred orientations in the active and inactive states with largely parallel orientation relative to the membrane of most of the helices. The distinct locations of the different isoforms, exposing them to different effectors and regulators, coupled with different G-domain-membrane orientation, suggests synergy between this type of recognition motif and the specificity conferred by the HVR, thereby validating the concept of isoform specificity in Ras.
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Abbreviations
- aa:
-
Amino acid
- AFM:
-
Atomic-force microscopy
- ATR-FTIR:
-
Attenuated total-reflection Fourier-transform infrared
- DMPC:
-
1,2-Dimyristoyl-sn-glycero-3-phosphocholine
- DOPC:
-
1,2-Dioleoyl-sn-glycero-3-phosphocholine
- DOPG:
-
1,2-Dioleoyl-sn-glycero-3-phospho-(1′-rac-glycerol)
- DPPC:
-
1,2-Dipalmitoyl-sn-glycero-3-phosphocholine
- DPPG:
-
1,2-Dipalmitoyl-sn-glycero-3-phospho-(1′-rac-glycerol)
- HVR:
-
Hypervariable region
- IRRAS:
-
Infrared reflection absorption spectroscopy
- ld :
-
Liquid-disordered
- lo :
-
Liquid-ordered
- MD:
-
Molecular dynamics
- PM:
-
Plasma membrane
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Acknowledgments
This research was supported by the Deutsche Forschungsgemeinschaft (DFG, SFB 642) and the International Max Planck Research School (IMPRS). We are grateful to Christine Nowak for technical assistance. Furthermore, we thank Dr Andreas Kerth, Dr Annette Meister, and Professor Alfred Blume (all Martin-Luther-Universität Halle-Wittenberg, Germany) and Dr Andrea Gohlke for their help in setting up the IRRAS measurements in our laboratory. Finally, RW wants to thank Professor Dr Peter Laggner, to whom this work is dedicated, for many stimulating discussions in various fields of membrane biophysics over the years.
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Special Issue: Scattering techniques in biology—marking the contributions to the field of Peter Laggner, on the occasion of his 68th birthday.
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Kapoor, S., Weise, K., Erlkamp, M. et al. The role of G-domain orientation and nucleotide state on the Ras isoform-specific membrane interaction. Eur Biophys J 41, 801–813 (2012). https://doi.org/10.1007/s00249-012-0841-5
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DOI: https://doi.org/10.1007/s00249-012-0841-5