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. 2000 Apr 11;97(9):4724–4729. doi: 10.1073/pnas.080063997

Figure 4.

Figure 4

Functional comparison of yeast L143P and human L136P mutants. (A) Transformants of rrn3− strain RLY303 expressing polyoma-tagged wild-type yRrn3 (yWT), yeast mutant L143P (yL143P), wild-type hRrn3 (hWT), human mutant L136P (hL136P), or empty expression vector are compared. Colonies were streaked on glucose to assay rrn3 complementation or onto galactose as a positive control and were incubated at 30°C or 37°C for 3 days. (B) Five-fold serial dilutions of the yeast and hRrn3 transformants compared in A were spotted onto glucose and incubated for 3 days at 30°C or 37°C. (C) Western blot monitoring expression of Rrn3 proteins in RLY303 transformants. Crude lysates were prepared from cells grown to OD 600 = 2.0 at 30°C in galactose media lacking leucine to select for the expression construct. Equal amounts of protein were loaded in each lane, and the blot was probed with a monoclonal antibody against the polyoma epitope. The identity of each expression construct is indicated above the lanes, and the positions of molecular weight standards are indicated to the right of the figure.