Figure 2.

Recombinant SMAX1 Is Unstable in Protein Extracts from Arabidopsis Seedlings.

(A) to (C) Immunoblots of in vitro degradation time courses of SMAX1-His in wild-type Col-0 (A), kai2 (B), and max2 (C) mutant protein extracts. Heterologously expressed and purified SMAX1-His was added to soluble protein extracts from 7-d-old Arabidopsis seedlings to compose 10% of the total protein concentration. Aliquots (50 µg) of the mixture were incubated at 24°C and boiled for 10 min at the indicated time points. Protein (33 µg) from each boiled aliquot was separated by SDS-PAGE, blotted, and probed with polyclonal anti-His antibody. The boiled plant protein extract incubated with SMAX1-His for 60 min served as the control. Where indicated in (C), 40 µM MG-132 was added to protein extracts prior to addition of SMAX1-His.