Subcellular Fractionation and Proteomics of Nuclear Envelopes
互联网
352
Because of its many connections to other cell systems, the nuclear envelope (NE)is essentially impossible to purify to homogeneity.
To circumvent these problems, we developed a subtractive proteomics approach in which the fraction of interest and a fraction
known to contaminate the fraction of interest are separately analyzed, and proteins identified in both fractions are subtracted
from the data set. This requires that the contaminating fraction can be purified to homogeneity. In this case, microsomal
membranes (MMs) are used to represent endoplasmic reticulum contamination, allowing the identification of transmembrane proteins
specific to the NE. To circumvent problems commonly associated with analyzing membrane proteins, the multidimensional protein
identification technology (MudPIT) proteomics methodology is employed.
