A fiducial-assisted strategy compatible with resolving small MFS transporter structures in multiple conformations using cryo-EM.

ABSTRACT: Advancements in cryo-EM have stimulated a revolution in structural biology. Yet, for membrane proteins near the cryo-EM size threshold of approximately 40 kDa, including transporters and G-protein coupled receptors, the absence of distinguishable structural features makes image alignment and structure determination a significant challenge. Furthermore, resolving more than one protein conformation within a sample, a major advantage of cryo-EM, represents an even greater degree of difficulty. Here, we describe a strategy for introducing a rigid fiducial marker (BRIL domain) at the C-terminus of membrane transporters from the Major Facilitator Superfamily (MFS) with AlphaFold2. This approach involves fusion of the last transmembrane domain helix of the target protein with the first helix of B

SUBMITTER: Xie P 

PROVIDER: S-EPMC11695964 | biostudies-literature | 2025 Jan

REPOSITORIES: biostudies-literature