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. Author manuscript; available in PMC: 2006 Dec 19.
Published in final edited form as: J Biol Chem. 2003 Sep 22;278(48):48445–48452. doi: 10.1074/jbc.M309238200

Fig.4.

Fig.4

Atg23 functions at the stage of Cvt vesicle formation. A, precursor Ape1 is membrane-associated in atg23Δ cells. The P13 fraction from atg23Δ (KTY14) spheroplasts was collected and loaded on the bottom of a Ficoll step gradient in the presence or absence of the detergent Triton X-100 (Tx). After centrifugation, the float (F) fractions were collected and analyzed by immunoblot using anti-Ape1 antiserum as described under “Experimental Procedures.” Lysis conditions were verified by immunoblot analysis using anti-Pgk1. B, precursor Ape1 is protease-sensitive in atg23Δ pep4Δ cells. The 13,000 × g pellet (P13) fraction was collected from atg23Δ pep4Δ (KTY22) lysed spheroplasts, subjected to treatment with proteinase K in the presence or absence of Triton X-100 (Tx), and analyzed by immunoblot using anti-Ape1 antiserum as described under “Experimental Procedures.” Lysis conditions were verified by immunoblot analysis using anti-Pgk1 and anti-Pho8 antisera. T, total; S13, 13,000 × g supernatant fraction.