Fig. 2.

Comparison of the half-lives of PCNA DNA-sliding clamp protein and translation initiation factor eIF2α in Haloferax volcanii wild-type (WT, ○) and panA deletion (ΔpanA, •) strains. (a) ³⁵S-pulse-chase labeling was coupled with immunoprecipitation using anti-PCNA and anti-eIF2α polyclonal antibodies to monitor the respective half-lives of PCNA and eIF2α in wild-type and ΔpanA mutant strains over a 12-h time course (with 12 h results similar to 1 h, data not shown). These data are representative of at least three independent experiments. (b) A graphical representation of the PCNA- and eIF2α-specific protein band intensities in percentages relative to 0 min for each protein, were used to calculate protein half-lives. (c) Similar overall levels of ³⁵S-label were incorporated into the bulk-protein of wild-type (lanes 4–6) and ΔpanA (lanes 1–3) strains. Total cell protein was radiolabeled with ³⁵S-methionine and ³⁵S-cysteine, separated by 12% SDS-PAGE and analyzed by autoradiography as described in Materials and methods.