Detection of nonfunctional overexpression gene products using two‐dimensional gel electrophoresis with a narrowed pH range

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Abstract
A commonly used technique in the analysis of yeast protein function is the overexpression of cloned genes. In the case that overexpression does not lead to an altered phenotype a stable synthesis of the protein has to be demonstrated. Here an example is shown where overexpression of the yeast HYP2 genes, coding for a hypusine‐containing protein, was alternatively analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) alone or by two‐dimensional (2‐D) gel electrophoresis, using the narrowed pH range of 4.5–5.4. The results of the SDS‐PAGE suggested a stable overproduction of HYP gene product, whereas 2‐D analysis revealed an accumulation of nonfunctional protein isoforms.