Periodicity of Deoxyribonuclease I Digestion of Chromatin

Abstract

Two methods have been used to measure the single-strand lengths of the DNA fragments produced by deoxyribonuclease I digestion of chromatin. The average lengths obtained are muliples of about 10.4 bases, significantly different from the value of 10 previously reported. This periodicity in fragment lengths is closely related to the periodicity of the DNA double helix in chromatin, but the two values need not be exactly the same.

DNase II digestion of the nucleosome core: precise locations and relative exposures of sites

“…Digestion attacked by DNase I with those attacked by DNase II, it is necessary to assign absolute fragment lengths to the bands in the DNase II-produced pattern. Such an assignment was made for the DNase I-produced bands by coelectrophoresis with sequenced DNA fragments (7,10), s0 the DNase I-produced bands could now in turn be assigned sizes by coelectrophoresis with such a 'standardized' DNase I-produced pattern. However, DNase II produces a fragment with a 3' terminal phosphate (14) while DNase I produces a fragment with a 3' terminal hydroxyl (15) [ extra phosphate increased the mobility of the fragment by less than one half base, because if this were the case, one would be unlikely to assign a size one full base 'out of step'.…”

“…Fragment lengths were assigned to individual bands in both DNase I-and DNase I-produced patterns (see Figure 1) by electrophoresis in adjacent channels samples of end-labelled DNA from a DNase I digest of nuclei, a pattern whose bands have been assigned fragment lengths by coolectrophoresis with multiple sequenced restriction fragments (7,10). The unknown channel was then sized by matching bands at a position low enough in the gel where the lineup between channels was obvious (see also discussion relevant to Figure 3 below).…”

“…Bands were assigned fragment sizes by counting down from characteristic bands in the gol, specifically the band corresponding to the fragment 20 bases long in band 'B2' (see reference 10).…”

The helical periodicity of DNA on the nucleosome

“…Ironically, however, one of the quantities used in making the prediction, namely the screw of DNA on the nucleosome, was shortly thereafter called into question by more accurate measurements, made by ourselves and our colleagues, of the average distance between DNAse I cutting sites of the DNA in nuclei, which give a value of about 10.4 base pairs per turn, rather than 10 (7,8).…”

Sequence-dependent Nucleosome Structural and Dynamic Polymorphism. Potential Involvement of Histone H2B N-Terminal Tail Proximal Domain

“…Sequencing gels were prepared according to standard protocols, except that the acrylamide/bisacrylamide weight ratio was decreased from the usual 19:1 value to 12:1, to better resolve mixed-sequence fragments. (We failed to decrease that ratio to 6 -8:1, as described, 23,24 because the gel would not stick onto the glass plates. )…”