Conformational microheterogeneity in a DNA double helix: structure of restriction endonuclease Bam H1 recognition site
Sarma, M.H.; Dhingra, M.M.; Gupta, G.; Sarma, R.H.
Biochemical and Biophysical Research Communications 131(1): 269-276
1985
ISSN/ISBN: 0006-291X PMID: 2994650 Document Number: 256184
Structural studies using 500 MHz 1H NMR spectroscopy on BAM H1 recognition site d(GGATCC)2 in solution at 19.degree. is reported. The resonances from the sugar ring and base protons have been assigned from the 2D-COSY and NOESY spectra. Analyses of the NOESY cross-peaks between the base protons H8/H6 and sugar protons H2'/H2'', H3' reveal that the nucleotide units G2, A3 and C6 adopt (C3'-endo, .chi. = 240.degree.-260.degree.) conformation while G1, T4 and C5 exhibit (C2'-endo, .chi. = 240.degree.-260.degree.) conformation. NMR data clearly suggest that the two strands of d(GGATCC)2 are conformationally equivalent and there is a structural two-fold between the two A-T pairs. The above information and the NOESY data are used to generate a structural model of d(GGATCC)2. The important features are: (i) G1-G2 stack, the site of cleavage, shows an alteration in sugar pucker i.e. C2'-endo, C3'-endo as in a B-A junction, (ii) G2-A3 stack adopts a mini A-DNA, both the sugars being C3'-endo, (iii) A3-T4 stack, the site of two-fold, displays an A-B junction with alternation in sugar pucker as C3'-endo, C2'-endo, (iv) T4-C5 stack adopts a mini B-DNA both the sugars being C2'-endo and (v) C5-C6 stack exhibits a B-A junction with C2'-endo, C3'-endo sugar puckers. Thus, our studies demonstrate that conformational microheterogeneity with a structural two fold, is present in the Bam H1 recognition site.