Biomedical subjects

H Clausen-Schaumann

Publications and source records attributed to H Clausen-Schaumann.

6 recordsLinked to original sources

PubMed2000

Mechanical stability of single DNA molecules.

Using a modified atomic force microscope (AFM), individual double-stranded (ds) DNA molecules attached to an AFM tip and a gold surface were overstretched, and the mechanical stability of the DNA double helix was investigated. In lambda-phage DNA the previously reported B-S transition at 65 piconewtons (pN) is followed by a second conformational transition, during which the DNA double helix melts into two single strands. Unlike the B-S transition, the melting transition exhibits a pronounced force-loading-rate dependence and a marked hysteresis, characteristic of a nonequilibrium conformational transition. The kinetics of force-induced melting of the double helix, its reannealing kinetics, as well as the influence of ionic strength, temperature, and DNA sequence on the mechanical stability of the double helix were investigated. As expected, the DNA double helix is considerably destabilized under low salt buffer conditions (</=10 mM NaCl), while high ionic strength buffers (1 M NaCl) stabilize the double-helical conformation. The mechanical energy that can be deposited in the DNA double helix before force induced melting occurs was found to decrease with increasing temperature. This energy correlates with the base-pairing free enthalpy DeltaG(bp)(T) of DNA. Experiments with pure poly(dG-dC) and poly(dA-dT) DNA sequences again revealed a close correlation between the mechanical energies at which these sequences melt with base pairing free enthalpies DeltaG(bp)(sequence): while the melting transition occurs between 65 and 200 pN in lambda-phage DNA, depending on the loading rate, the melting transition is shifted to approximately 300 pN for poly(dG-dC) DNA, whereas poly(dA-dT) DNA melts at a force of 35 pN.

Bacteriophage lambda↗

PubMed2000

Force spectroscopy with single bio-molecules.

For many biological molecules, force is an important functional and structural parameter. With the rapidly growing knowledge about the relation between structure, function, and force, single-molecule force spectroscopy has become a versatile analytical tool for the structural and functional investigation of single bio-molecules in their native environments. Within the past year, detailed insights into binding potentials of receptor ligand pairs, protein folding pathways, molecular motors, DNA mechanics and the functioning of DNA-binding agents (such as proteins and drugs), as well as the function of molecular motors, have been obtained.

DNA↗

PubMed1999

How strong is a covalent bond?

The rupture force of single covalent bonds under an external load was measured with an atomic force microscope (AFM). Single polysaccharide molecules were covalently anchored between a surface and an AFM tip and then stretched until they became detached. By using different surface chemistries for the attachment, it was found that the silicon-carbon bond ruptured at 2.0 +/- 0.3 nanonewtons, whereas the sulfur-gold anchor ruptured at 1.4 +/- 0.3 nanonewtons at force-loading rates of 10 nanonewtons per second. Bond rupture probability calculations that were based on density functional theory corroborate the measured values.

Journal Article↗

PubMed1999

Sequence-dependent mechanics of single DNA molecules.

Atomic force microscope-based single-molecule force spectroscopy was employed to measure sequence-dependent mechanical properties of DNA by stretching individual DNA double strands attached between a gold surface and an AFM tip. We discovered that in lambda-phage DNA the previously reported B-S transition, where 'S' represents an overstretched conformation, at 65 pN is followed by a nonequilibrium melting transition at 150 pN. During this transition the DNA is split into single strands that fully recombine upon relaxation. The sequence dependence was investigated in comparative studies with poly(dG-dC) and poly(dA-dT) DNA. Both the B-S and the melting transition occur at significantly lower forces in poly(dA-dT) compared to poly(dG-dC). We made use of the melting transition to prepare single poly(dG-dC) and poly(dA-dT) DNA strands that upon relaxation reannealed into hairpins as a result of their self-complementary sequence. The unzipping of these hairpins directly revealed the base pair-unbinding forces for G-C to be 20 +/- 3 pN and for A-T to be 9 +/- 3 pN.

Bacteriophage lambda↗

PubMed1998

Atomic force microscope imaging of phospholipid bilayer degradation by phospholipase A2.

We have investigated the time course of the degradation of a supported dipalmitoylphosphatidylcholine bilayer by phospholipase A2 in aqueous buffer with an atomic force microscope. Contact mode imaging allows visualization of enzyme activity on the substrate with a lateral resolution of less than 10 nm. Detailed analysis of the micrographs reveals a dependence of enzyme activity on the phospholipid organization and orientation in the bilayer. These experiments suggest that it is possible to observe single enzymes at work in small channels, which are created by the hydrolysis of membrane phospholipids. Indeed, the measured rate of hydrolysis of phospholipids corresponds very well with the enzyme activity found in kinetic studies. It was also possible to correlate the number of enzymes at the surface, as calculated from the binding constant to the number of starting points of the hydrolysis. In addition, the width of the channels was found to be comparable to the diameter of a single phospholipase A2 and thus further supports the single-enzyme hypothesis.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1998

Direct detection of domains in phospholipid bilayers by grazing incidence diffraction of neutrons and atomic force microscopy.

The geometry of domains in phospholipid bilayers of binary (1:1) mixtures of synthetic lecithins with a difference in chain length of four methylene groups has been studied by two independent, direct and complementary methods. Grazing incidence diffraction of neutrons provided gel domain sizes of less than 10 nm in both the gel and the coexistence phase of the mixture, while no domains were detected for the fluid phase. For the coexistence region, the neutron data suggest that domains grow in number rather than in size with decreasing temperature. Atomic force microscopy was used to study gel phase size and shape of the domains. The domains imaged by atomic force microscopy exhibit a rather irregular shape with an average size of 10 nm, thus confirming the neutron results for this phase. The good agreement between atomic force microscopy and neutron results, despite the completely different nature of their observables, has potential for the future development of refined models for the interpretation of neutron data from heterogeneous membranes in terms of regularly spaced and spatially extended scatterers.

Calorimetry, Differential Scanning↗