Production of mitotic regulators using an autoselection system for protein expression in budding yeast.
A novel protein expression system in budding yeast is described which has been used to express many yeast mitotic regulators as well as a wide range of other recombinant proteins from several different species. The expression system relies on autoselection with essential genes to maintain high copy numbers of expression plasmids. Autoselection permits expression cells to be grown in rich medium with no need for plasmid selection with drugs or nutritional conditions. This optimizes growth and expression of recombinant proteins. The use of the expression system is illustrated by purifying budding yeast mitotic regulators, Cdc14 and Net1, and recapitulating their activities in vitro.
Geymonat
Marco
M
Stem Cell Biology and Developmental Genetics, National Institute for Medical Research, London, UK.
Spanos
Adonis
A
Sedgwick
Steven
S
D000595:Amino Acid Sequence; D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000096:biosynthesis*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000096:biosynthesis; D010957:Plasmids; D017027:Protein Tyrosine Phosphatases / Q000096:biosynthesis*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis*
C074860:CDC14 protein, S cerevisiae; D018797:Cell Cycle Proteins; C119300:Net1 protein, S cerevisiae; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; D017027:Protein Tyrosine Phosphatases