The Self‐Association of the KRAS4b Protein is Altered by Lipid‐Bilayer Composition and Electrostatics

• Published in: Angewandte Chemie International Edition 2023-04, Vol.62 (18), p.e202218698-n/a
• Main Authors: Lee, Ki‐Young, Ikura, Mitsuhiko, Marshall, Christopher B.
• Format: Article
• Language: English
• Subjects: Charge reversal; Composition; Dimerization; Dimers; Electrostatic properties; Electrostatics; Interfaces; K-Ras protein; KRAS Dimers; Lipid Bilayers - chemistry; Lipid composition; Lipids; Membrane proteins; Membranes; Molecular Conformation; Nanodiscs; NMR; NMR Spectroscopy; Nuclear magnetic resonance; Phosphatidylserine; Plastic Interactions; Proteins; Proteins - metabolism; Proto-Oncogene Proteins p21(ras) - genetics; Proto-Oncogene Proteins p21(ras) - metabolism; Raf protein; Self-assembly; Signaling; Static Electricity
• ISSN: 1433-7851; 1521-3773
• DOI: 10.1002/anie.202218698

KRAS is a peripheral membrane protein that regulates multiple signaling pathways, and is mutated in ≈30 % of cancers. Transient self‐association of KRAS is essential for activation of the downstream effector RAF and oncogenicity. The presence of anionic phosphatidylserine (PS) lipids in the membrane was shown to promote KRAS self‐assembly, however, the structural mechanisms remain elusive. Here, we employed nanodisc bilayers of defined lipid compositions, and probed the impact of PS concentration on KRAS self‐association. Paramagnetic NMR experiments demonstrated the existence of two transient dimer conformations involving alternate electrostatic contacts between R135 and either D153 or E168 on the “α4/5‐α4/5” interface, and revealed that lipid composition and salt modulate their dynamic equilibrium. These dimer interfaces were validated by charge‐reversal mutants. This plasticity demonstrates how the dynamic KRAS dimerization interface responds to the environment, and likely extends to the assembly of other signaling complexes on the membrane. Self‐association of the peripheral membrane protein KRAS4b promotes oncogenic signaling. Using nanodiscs to enable paramagnetic relaxation enhancement NMR experiments, we show that KRAS4b dimers associate through distinct conformations depending on lipid composition. Both states involve the α4‐α5 interface, however increasing phosphatidylserine concentration alters the relative protomer orientation, shifting R135 electrostatic interactions from D153 to E168.