Characterization of Postmortem Human Brain Proteins by Two‐Dimensional Gel Electrophoresis

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Abstract
The proteins of membrane and cytosol fractions from frozen human postmortem brain were analyzed by 2-dimensional gel electrophoresis (isoelectric range: 5.1-6.0) and both Coomassie-blue and ammoniacal Ag staining. Cytosol preparations were analyzed from 6 different postmortem brains from patients with various neurologic diagnoses and immediate causes of death. Intervals between death and brain freezing (-70.degree. C) ranged from 2-20 h. The vast majority of proteins detected in these cytosol fractions had identical MW and isoelectric points in each of 6 human brains examined. In some tissue samples, tubulin was either quantitatively decreased or undetectable. The possibility that this partial or complete depletion of tubulin was related to postmortem interval and/or brain freezing was studied with rat forebrain tissue. Rat brain incubated at room temperature for up to 24 h did not reproduce the changes seen in the region of human cytosol tubulin. Other changes seen in the 2-dimensional electrophoretic pattern of rat cytosol proteins did relate to postmortem interval, brain freezing, or both. Rough endoplasmic reticulum (RER) and smooth endoplasmic reticulum were prepared from 3 human brains, with highly reproducible 2-dimensional patterns. Protein analysis of these membrane fractions revealed that human RER contained significant amounts of tubulin, in contrast to rat RER which contained no detectable tubulin. This discrepancy was elucidated by allowing rat brains to remain at room temperature for 24 h before freezing; gels of rat RER prepared from this tissue showed that tubulin subunits were present.