Silatrane-based surface chemistry for immobilization of DNA, protein-DNA complexes and other biological materials.

The procedure of surface functionalization based on the use of 1-(3-Aminopropyl)silatrane (APS) instead of our early procedure utilizing aminopropyl triethoxy silane (APTES) is described. Unlike APTES, APS is less reactive and extremely resistant to hydrolysis and polymerization at neutral pH. The kinetics of DNA adsorption to APS-mica was studied. The results are consistent with a diffusion controlled mechanism suggesting that DNA molecules bind irreversibly with the surface upon immobilization. This conclusion is supported by the data on imaging of supercoiled DNA, the labile conformations of which are very sensitive to the conditions at the surface-liquid interface. In addition, we demonstrated directly that the segments of DNA molecules could move along the surface if the sample is imaged in aqueous solution without drying of the sample. Using the time-lapse mode of AFM imaging we visualized the transition of purine-pyrimidine sequence in supercoiled DNA from intramolecular triple-helical conformation (H-form) into the B-helix upon the change of pH from acidic (pH 5) to neutral. The mechanisms of the H-to-B transitions and the correlation of the local structural transitions with a global DNA conformation are discussed.

[1]  A. Rich,et al.  The Zalpha domain from human ADAR1 binds to the Z-DNA conformer of many different sequences. , 1998, Nucleic acids research.

[2]  C. Bustamante,et al.  Visualizing protein-nucleic acid interactions on a large scale with the scanning force microscope. , 1996, Annual review of biophysics and biomolecular structure.

[3]  C. Bustamante,et al.  Scanning force microscopy of DNA deposited onto mica: equilibration versus kinetic trapping studied by statistical polymer chain analysis. , 1996, Journal of molecular biology.

[4]  Y. Lyubchenko,et al.  Atomic force microscopy of DNA and protein-DNA complexes using functionalized mica substrates. , 2001, Methods in molecular biology.

[5]  H. Hansma,et al.  Properties of biomolecules measured from atomic force microscope images: a review. , 1997, Journal of structural biology.

[6]  P. Hough,et al.  Recent advances in atomic force microscopy of DNA. , 1993, Scanning.

[7]  C. Siegerist,et al.  Reproducible Imaging and Dissection of Plasmid DNA Under Liquid with the Atomic Force Microscope , 1992, Science.

[8]  D. Lohr,et al.  Evidence for nonrandom behavior in 208-12 subsaturated nucleosomal array populations analyzed by AFM. , 1999, Biochemistry.

[9]  Z. Shao,et al.  High‐resolution atomic‐force microscopy of DNA: the pitch of the double helix , 1995, FEBS letters.

[10]  J. Griffith,et al.  DNA electron microscopy. , 1981, CRC critical reviews in biochemistry.

[11]  Z. Shao,et al.  Atomic force microscopy of DNA molecules , 1992, FEBS letters.

[12]  Wigbert J. Siekhaus,et al.  Atomic force microscopy of mammalian sperm chromatin , 1993, Chromosoma.

[13]  Y. Lyubchenko,et al.  Atomic force microscopy imaging of double stranded DNA and RNA. , 1992, Journal of biomolecular structure & dynamics.

[14]  D. Keller,et al.  Scanning force microscopy under aqueous solutions. , 1997, Current opinion in structural biology.

[15]  Y. Lyubchenko,et al.  Atomic force microscopy: a powerful tool to observe biomolecules at work. , 1999, Trends in cell biology.

[16]  Y. Lyubchenko,et al.  Atomic force microscopy of long DNA: imaging in air and under water. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[17]  Wolfgang M. Heckl,et al.  Procedures in scanning probe microscopy , 1998 .

[18]  C. Bustamante,et al.  Circular DNA molecules imaged in air by scanning force microscopy. , 1992, Biochemistry.

[19]  M. Hegner,et al.  Immobilizing DNA on gold via thiol modification for atomic force microscopy imaging in buffer solutions , 1993, FEBS letters.

[20]  R. Sinden,et al.  The structure of intramolecular triplex DNA: atomic force microscopy study. , 2001, Journal of molecular biology.

[21]  Richard R. Sinden,et al.  DNA structural transitions within the PKD1 gene , 1999, Nucleic Acids Res..

[22]  Y. Lyubchenko,et al.  Visualization of supercoiled DNA with atomic force microscopy in situ. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[23]  C. Bustamante,et al.  Determination of heat‐shock transcription factor 2 stoichiometry at looped DNA complexes using scanning force microscopy. , 1995, The EMBO journal.

[24]  Y. Lyubchenko,et al.  Atomic force microscopy imaging of DNA covalently immobilized on a functionalized mica substrate. , 1999, Biophysical journal.

[25]  C. Bustamante,et al.  Visualization and analysis of chromatin by scanning force microscopy. , 1997, Methods.

[26]  R. Sinden,et al.  Structure and dynamics of supercoil-stabilized DNA cruciforms. , 1998, Journal of molecular biology.

[27]  H. Hansma,et al.  Applications for atomic force microscopy of DNA. , 1995, Biophysical journal.

[28]  N R Cozzarelli,et al.  Conformational and thermodynamic properties of supercoiled DNA. , 1994, Annual review of biophysics and biomolecular structure.

[29]  H. Hansma,et al.  Atomic force microscopy and other scanning probe microscopies. , 1998, Current opinion in chemical biology.

[30]  R. Sinden,et al.  A cruciform structural transition provides a molecular switch for chromosome structure and dynamics. , 2000, Journal of molecular biology.

[31]  R. Sinden,et al.  Structure and dynamics of three-way DNA junctions: atomic force microscopy studies. , 2000, Nucleic acids research.