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Measurement of the differences in electrophoretic mobilities of individual molecules of E. coli β‐galactosidase provides insight into structural differences which underlie enzyme microheterogeneity
Author(s) -
Nichols Ellert R.,
Craig Douglas B.
Publication year - 2008
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200800060
Subject(s) - electrophoresis , capillary electrophoresis , chemistry , molecule , beta galactosidase , electrophoretic mobility shift assay , enzyme , polymer , resolution (logic) , chromatography , biochemistry , escherichia coli , organic chemistry , gene expression , gene , artificial intelligence , computer science
The electrophoretic mobility and catalytic activity of individual molecules of Escherichia coli β‐galactosidase were measured using CE‐LIF detection. Both the mobility and activity were reproducible for each molecule but differed between individual molecules. Assays were performed using uncoated capillaries and capillaries coated with different polymers, using enzymes from different sources and by three different experimental protocols. In all cases the observed ranges in electrophoretic mobilities were similar. The observed range in the electrophoretic mobility may be explained by structural microheterogeneity resulting in a gain or loss of up to 1.6 suppressed charge units. There was no observed relationship between the observed activities and electrophoretic mobilities. If the finding that individual β‐galactosidase molecules have heterogeneous electrophoretic mobility can be extended to other proteins, this may limit the resolution possible for capillary zone electrophoresis protein separations.