NLA Forms Homodimers via Its RING Finger Domain.
(A) Schematic diagrams of full-length NLA and its deletion derivatives used to test for NLA homodimerization. MBP or MBP-NLA was used as a bait protein and GST-NLA (amino acids 1 to 335), GST-SPX (amino acids 1 to 270), or GST-RING (amino acids 220 to 335) was used as a prey protein.
(B) In vitro pull-down assay of MBP-NLA and GST-NLA. Purified GST-NLA protein as prey was mixed with MBP or MBP-NLA as bait.
(C) In vitro pull-down assay of MBP-NLA and GST-SPX.
(D) In vitro pull-down assay of MBP-NLA and GST-RING. For (B) to (D), the reaction mix contained 1 μg each of prey and bait proteins. Pulled-down protein was analyzed by protein gel blot using anti-GST antibody.