A sampling of the yeast proteome

Abstract

In this study, we examined yeast proteins by two-dimensional (2D) gel electrophoresis and gathered quantitative information from about 1,400 spots. We found that there is an enormous range of protein abundance and, for identified spots, a good correlation between protein abundance, mRNA abundance, and codon bias. For each molecule of well-translated mRNA, there were about 4,000 molecules of protein. The relative abundance of proteins was measured in glucose and ethanol media. Protein turnover was examined and found to be insignificant for abundant proteins. Some phosphoproteins were identified. The behavior of proteins in differential centrifugation experiments was examined. Such experiments with 2D gels can give a global view of the yeast proteome.

FIG. 1

2D gels. The horizontal axis is the isoelectric focusing dimension, which stretches from pH 6.7 (left) to pH 4.3 (right). The vertical axis is the polyacrylamide gel dimension, which stretches from about 15 kDa (bottom) to at least 130 kDa (top). For panel A, extract was made from cells in log phase in glucose; for panel B, cells were grown in ethanol. The spots labeled 1 through 6 are unidentified proteins highly induced in ethanol.

FIG. 2

Correlation of protein abundance with adjusted mRNA abundance. The number of molecules per cell of each protein is plotted against the number of molecules per cell of the cognate mRNA, with an r_p of 0.76. Note the logarithmic axes. Data for mRNA were taken from references and and combined as described in Materials and Methods.

FIG. 3

Correlation of protein abundance with CAI. The number of molecules per cell of each protein is plotted against the CAI for that protein. Note the logarithmic scale on the protein axis. Data for the CAI are from the YPD database (13).

FIG. 4

Distribution of CAI over the whole genome, shown in intervals of 0.030 (i.e., there are 150 genes with a CAI between 0.000 and 0.030, inclusive; 31 genes with a CAI between 0.031 and 0.060; 269 genes with a CAI between 0.061 and 0.090; 1,296 genes with a CAI between 0.091 and 0.120; etc.). The distribution peaks with 2,028 genes with a CAI between 0.121 and 0.150.

FIG. 5

Phosphorylated proteins. (A) Mixture of ³²P-labeled proteins and ³⁵S-labeled proteins. Two separate labeling reactions were done, one with ³²P and one with ³⁵S, and extracts were mixed and run on a 2D gel. Spots marked with numbers rather than gene names represent spots noted on ³⁵S gels but unidentified. Spots labeling with ³²P were identified by (i) increased labeling compared to the ³⁵S-only gel (not shown); (ii) the characteristic fuzziness of a ³²P-labeled spot; and (iii) the decay of signal intensity seen on exposures made 4 weeks later (not shown). A minor form of Tpi1 and at least six minor forms of Tif1 have been noted in overexpression experiments (see also Fig. 6B); positions of the minor forms are indicated by circles. (B) ³²P-only labeling. The major form of Tpi1, which is not labeled with ³²P, is indicated by a large circle; positions of seven forms of Tif1 are indicated by smaller circles.

FIG. 6

Fractionation by centrifugation. (A) Proteins in the supernatant of a 100,000 × g, 30-min spin; proteins in the pellet of a 16,000 × g, 10-min spin. Supernatant fractions examined in multiple experiments done over a wide range of g forces looked similar to each other, as did the pellet fractions.