Helical repeat and chirality effects on DNA gel electrophoretic mobility

Abstract

We determined a value of 10.34 +/- 0.04 base pairs (bp) per turn for the helical repeat of bent DNA sequences of the form A6N4-A6N5 by estimating the sequence repeat required to produce a planar curve, as judged from the maximum in the electrophoretic mobility anomaly of multimers containing different sequence repeats (10.00, 10.33, 10.50, 10.67, and 11.00 bp per turn). This result provides the basis for a method to evaluate the helical repeat of any DNA segment by comparative electrophoresis measurements. The sequence of interest is placed between two A-tract bends and the phasing is varied over an entire helical turn. Knowledge of the number of base pairs between the bends in the cis isomer, which has the lowest electrophoretic mobility, allows calculation of the average helical repeat of the inserted sequence. In the course of these experiments we observed an unexpected dependence of electrophoretic mobility on the shape of DNA molecules: in high-percentage polyacrylamide gels, those bent molecules for which we deduced a right-handed superhelical form are less retarded than their homologous left-handed isomers. To explain this finding we propose that superhelical chirality influences the choice of DNA migration pathway, leading to rotation of the DNA molecule relative to the local coordinate frame in the gel. High-percentage gels have sufficiently close contact with the right-handed DNA helical twist to differentiate the frictional consequences of right- and left-handed twisting motions.