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. 2006 Oct 23;175(2):249–259. doi: 10.1083/jcb.200605162

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Copyright © 2006, The Rockefeller University Press

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Figure 3. — Int-atTic40 is an intermediate in the import and membrane targeting of mature atTic40. (A) [³⁵S]pre-atTic40 was incubated with isolated chloroplasts in the presence of 100 μM ATP to promote formation of an early import intermediate. The chloroplasts were isolated and incubated in the presence of 2 mM ATP to promote import of the bound pre-atTic40 for the times indicated (chase). The chloroplasts were separated into membrane and soluble fractions by osmotic lysis and analyzed by SDS-PAGE and phosphorimaging. (B) Quantitative analysis of the total amounts of [³⁵S]-labeled pre-atTic40, int-atTic40, and mature atTic40 in the combined soluble and membrane fractions from A. (C) Samples from the same import experiment in A, but the intact chloroplasts were treated with 100 μg/ml thermolysin before separation into membrane and soluble fractions. (D) Quantitative analysis of the percentage of protease-resistant int-atTic40 or mature atTic40 (from C) relative to the total amount of each species (from A) at each time point in the import experiment. The samples in A and C were generated from the same import experiment and were analyzed on the same SDS-PAGE gel.