Analysis of DNA-HMG2 complex using AFM

Masato Saito, Yuji Murakami, Kenji Yokoyama, Shin-ichi Tate and Eiichi Tamiya

School of materials science, Japan Advanced Institute of Science and Technology (JAIST)

HMG2 constitutes a highly abundant class of non-histone chromosomal protein. HMG2 binds to DNA non-specifically and bends DNA. We attempted to visualize DNA-HMG2 complexes by using AFM. Different concentration of HMG2 such as 16-160 moles were mixed with 1 mole of 519 bp DNA amplified by PCR in 10 mM Tris-HCl (pH 8.0) buffer. The complexes were placed on a mica modified with g-aminopropyltriethoxysilane (g-APTES). AFM image of complex at high concentrate HMG2 (160 moles) was observed as condensed structure. However, when HMG2 concentration was reduced, complexes observed from unwinding structure to straight form. This indicated that HMG2 bends DNA by binding non-specifically at high concentration of HMG2. For measurement of DNA bending angle, we optimized the condition to 1:160 molar ratio in 60 mM NaCl, 10 mM Tris-HCl. As a result, the complex bending angle was 100.1Å}33.7Åã( meanÅ}SD ).