135 research outputs found
Analysis of Diffusion of Ras2 in Saccharomyces cerevisiae Using Fluorescence Recovery after Photobleaching
Binding, lateral diffusion and exchange are fundamental dynamic processes
involved in protein association with cellular membranes. In this study, we
developed numerical simulations of lateral diffusion and exchange of
fluorophores in membranes with arbitrary bleach geometry and exchange of the
membrane localized fluorophore with the cytosol during Fluorescence Recovery
after Photobleaching (FRAP) experiments. The model simulations were used to
design FRAP experiments with varying bleach region sizes on plasma-membrane
localized wild type GFP-Ras2 with a dual lipid anchor and mutant GFP-Ras2C318S
with a single lipid anchor in live yeast cells to investigate diffusional
mobility and the presence of any exchange processes operating in the time scale
of our experiments. Model parameters estimated using data from FRAP experiments
with a 1 micron x 1 micron bleach region-of-interest (ROI) and a 0.5 micron x
0.5 micron bleach ROI showed that GFP-Ras2, single or dual lipid modified,
diffuses as single species with no evidence of exchange with a cytoplasmic
pool. This is the first report of Ras2 mobility in yeast plasma membrane. The
methods developed in this study are generally applicable for studying diffusion
and exchange of membrane associated fluorophores using FRAP on commercial
confocal laser scanning microscopes.Comment: Accepted for publication in Physical Biology (2010). 28 pages, 7
figures, 3 table
Optimal design of multioutput sampling schedules - Software and applications to endocrine-metabolic and pharmacokinetic models
This paper describes a program for computing optimal sampling
schedules for multiinput-multioutput experiments designed for parameter
estimation of physiological systems models. Theory of the algorithm
and details of its implementation are given. Practical applications
of the software to models of glucose-insulin regulation, ketone body, and
insulin kinetics are presented. Results document the potentiality of the
software for designing experiments, and show that optimal design can
considerably reduce the number of samples withdrawn from a patient in
in vivo clinical studies
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