Figure 3.

 Clp-dependent turnover in vivo of λN-AA and λN-L1-AA. (A) Western blot analysis of λN-AA in wild-type (SG22163) or clpP mutant (SG22175) cells after spectinomycin blocking of protein synthesis. Cells were grown to mid-log phase at 32°C and induced with IPTG for 30 min before addition of spectinomycin. Samples were removed at the times shown into cold TCA, precipitated, and resuspended for electrophoresis in 15% SDS–polyacrylamide gels. Gels were blotted and probed with anti-λ-repressor antibody. (B) Turnover of λN-L1-AA in wild-type and clp mutants. Experimental conditions were the same as in A except 10%–20% tricine SDS–polyacrylamide gels were used, and Western blots were probed with the M2 anti-FLAG antibody and quantified by the Eagle Eye II gel imaging system. Curves and strains are marked as for Fig. 2.