FIG. 3.
Pulse-chase analysis of HemA turnover in adapted hemL mutant cells. A hemL deletion mutant of S. typhimurium (TE472) was grown in MOPS glycerol medium to an OD600 of 0.4 in the presence of 2 μM ALA (top panel in panel A; heme unlimited) or adapted to growth in the same medium but without ALA and grown to an OD600 of 0.4 (bottom panel in panel A; heme limited). Cultures were labeled and analyzed as described in the legend to Fig. 2, except that the chase with unlabeled methionine and cystine was extended as shown above each lane. Identical gels (not treated with fluor) were analyzed by using a PhosphorImager and its ImageQuant software to produce the data plotted (B). The calculated half-life of HemA protein in unlimited cells is ≈20 min (closed circles), compared with a half-life estimated to be in excess of 300 min in adapted, heme-limited cells (open circles).