Electrophoretic Heterogeneity of Grapevine Histones

摘要

Histones from grapevine leaves were isolated using classical extraction procedures. Different electrophoretic processes were tested to study the heterogeneity of grapevine histones. Polyacrylamide gels containing sodium dodecyl sulfate were useful for the fractionation of grapevine histone H1, H3, and H4 classes but inefficient for the isolation of histone H2A and H2B classes. Gels containing Triton X-100, used for the second dimension in two-dimensional electrophoresis, proved to be more efficient for the separation of all histone classes, as well as for that of their subfractions (structural variants and/or chemical modified forms) and resulted in optimum fractionation of grapevine nucleosomal proteins. The core histone H2B fraction, which is very sensitive to oxidation, seems to consist of a large number of subfractions, forming three sets of proteins with different mobility in polyacrylamide gels containing acetic acid, urea, and Triton X-100 (AUT-PAGE). Histones H1, H2A, and H3 also display internal heterogeneities with three forms for histone H1, four forms for histone H2A, and two forms for histone H3. In an attempt to define which of the Triton X-100 resolved subfractions represents oxidation forms, experimental oxidation of grapevine histones was performed.