Figure 1.

Purification of LAT1–CD98hc. (a) Immunoblot using anti-LAT1 polyclonal antibody under nonreducing conditions of HEK293 cells overexpressing LAT1 that were lysed in the absence (lane 1) or presence (lane 2) of 1 mM sodium aurothiomalate. The two bands that are visible are consistent with heterodimeric (123 kDa) and monomeric (55 kDa) LAT1. (b) Size-exclusion chromatogram of LAT1–CD98hc after purification by immunoprecipitation on a Superdex 200 10/300 column at 0.4 ml min⁻¹. The peak at a retention volume of 10.5 ml was used for cryo-EM. (c) Coomassie-stained SDS–PAGE gel of purified LAT1–CD98hc (lane 2) run under nonreducing conditions. (d) Coomassie-stained SDS–PAGE gel of purified LAT1–CD98hc (lane 2) run under reducing conditions and immunoblotted using anti-CD98hc monoclonal antibody (lane 3). Molecular-weight markers are labelled on the left in kDa.