Half-life of yeGFP-ssrA with expressed E. coli ClpXP protease

Range 22-91 min
Organism Budding yeast Saccharomyces cerevisiae
Reference Grilly C, Stricker J, Pang WL, Bennett MR, Hasty J. A synthetic gene network for tuning protein degradation in Saccharomyces cerevisiae. Mol Syst Biol. 2007 3: 127. abstract & p.3 right column 2nd paragraphPubMed ID17667949
Method Abstract: "Here, [investigators] use components of the Escherichia coli degradation machinery to construct a Saccharomyces cerevisiae strain that allows for tunable degradation of a tagged protein. Using a microfluidic platform tailored for single-cell fluorescence measurements, [they] monitor protein decay rates after repression using an ssrA-tagged fluorescent reporter." P.2 left column bottom paragraph: "[Investigators] have constructed a S. cerevisiae strain (CGD699) that allows tunable degradation of a tagged protein. To accomplish this, [they] expressed a modified E. coli ClpXP protease in yeast under the control of a repressible promoter."
Comments Abstract: "[Investigators] observe a half-life ranging from 91 to 22 min, depending on the level of activation of the degradation genes." P.3 right column 2nd paragraph: "Previous studies have used exponential fits to characterize the half‐life of the reporter. [Investigators] found that modeling decay as arising from a set of enzymatic Michaelis–Menten reactions led to excellent agreement between model and experiment (see below). However, in order to systematically compare with the previous fluorescent reporter degradation studies, [they] chose to first analyze the fluorescence trajectories with exponential fits that were reasonably accurate (Figure 3B). From these fits, [they] were able to calculate the mean half‐life for each concentration of IPTG [Isopropyl β-D-1-thiogalactopyranoside], as shown in Figure 3C. The half‐life decreases from a value of 91 min for no IPTG to 22 min for media containing 5 mM IPTG."
Entered by Uri M
ID 105190