Laura Finzi

Bio: Laura Finzi is an academic researcher from Emory University. The author has contributed to research in topics: DNA & DNA supercoil. The author has an hindex of 24, co-authored 79 publications receiving 3377 citations. Previous affiliations of Laura Finzi include University of Pennsylvania & University of New Mexico.

Direct mechanical measurements of the elasticity of single DNA molecules by using magnetic beads

Abstract: Single DNA molecules were chemically attached by one end to a glass surface and by their other end to a magnetic bead. Equilibrium positions of the beads were observed in an optical microscope while the beads were acted on by known magnetic and hydrodynamic forces. Extension versus force curves were obtained for individual DNA molecules at three different salt concentrations with forces between 10(-14) and 10(-11) newtons. Deviations from the force curves predicted by the freely jointed chain model suggest that DNA has significant local curvature in solution. Ethidium bromide and 4',6-diamidino-2-phenylindole had little effect on the elastic response of the molecules, but their extent of intercalation was directly measured. Conversely, the effect of bend-inducing cis-diamminedichloroplatinum (II) was large and supports the hypothesis of natural curvature in DNA.

Supercoiling and denaturation in Gal repressor/heat unstable nucleoid protein (HU)-mediated DNA looping.

Abstract: The overall topology of DNA profoundly influences the regulation of transcription and is determined by DNA flexibility as well as the binding of proteins that induce DNA torsion, distortion, and/or looping. Gal repressor (GalR) is thought to repress transcription from the two promoters of the gal operon of Escherichia coli by forming a DNA loop of ≈40 nm of DNA that encompasses the promoters. Associated evidence of a topological regulatory mechanism of the transcription repression is the requirement for a supercoiled DNA template and the histone-like heat unstable nucleoid protein (HU). By using single-molecule manipulations to generate and finely tune tension in DNA molecules, we directly detected GalR/HU-mediated DNA looping and characterized its kinetics, thermodynamics, and supercoiling dependence. The factors required for gal DNA looping in single-molecule experiments (HU, GalR and DNA supercoiling) correspond exactly to those necessary for gal repression observed both in vitro and in vivo. Our single-molecule experiments revealed that negatively supercoiled DNA, under slight tension, denatured to facilitate GalR/HU-mediated DNA loop formation. Such topological intermediates may operate similarly in other multiprotein complexes of transcription, replication, and recombination.

Tethered particle motion as a diagnostic of DNA tether length.

Abstract: The tethered particle motion (TPM) technique involves an analysis of the Brownian motion of a bead tethered to a slide by a single DNA molecule. We describe an improved experimental protocol with which to form the tethers, an algorithm for analyzing bead motion visualized using differential interference contrast microscopy, and a physical model with which we have successfully simulated such DNA tethers. Both experiment and theory show that the statistics of the bead motion are quite different from those of a free semiflexible polymer. Our experimental data for chain extension versus tether length fit our model over a range of tether lengths from 109 to 3477 base pairs, using a value for the DNA persistence length that is consistent with those obtained under similar solution conditions by other methods. Moreover, we present the first experimental determination of the full probability distribution function of bead displacements and find excellent agreement with our theoretical prediction. Our results show that TPM is a useful tool for monitoring large conformational changes such as DNA looping.

BASIC PENTACYSTEINE1, a GA Binding Protein That Induces Conformational Changes in the Regulatory Region of the Homeotic Arabidopsis Gene SEEDSTICK

Abstract: The mechanisms for the regulation of homeotic genes are poorly understood in most organisms, including plants. We identified BASIC PENTACYSTEINE1 (BPC1) as a regulator of the homeotic Arabidopsis thaliana gene SEEDSTICK (STK), which controls ovule identity, and characterized its mechanism of action. A combination of tethered particle motion analysis and electromobility shift assays revealed that BPC1 is able to induce conformational changes by cooperative binding to purine-rich elements present in the STK regulatory sequence. Analysis of STK expression in the bpc1 mutant showed that STK is upregulated. Our results give insight into the regulation of gene expression in plants and provide the basis for further studies to understand the mechanisms that control ovule identity in Arabidopsis.

Microfluidics: Fluid physics at the nanoliter scale

Abstract: Microfabricated integrated circuits revolutionized computation by vastly reducing the space, labor, and time required for calculations. Microfluidic systems hold similar promise for the large-scale automation of chemistry and biology, suggesting the possibility of numerous experiments performed rapidly and in parallel, while consuming little reagent. While it is too early to tell whether such a vision will be realized, significant progress has been achieved, and various applications of significant scientific and practical interest have been developed. Here a review of the physics of small volumes (nanoliters) of fluids is presented, as parametrized by a series of dimensionless numbers expressing the relative importance of various physical phenomena. Specifically, this review explores the Reynolds number Re, addressing inertial effects; the Peclet number Pe, which concerns convective and diffusive transport; the capillary number Ca expressing the importance of interfacial tension; the Deborah, Weissenberg, and elasticity numbers De, Wi, and El, describing elastic effects due to deformable microstructural elements like polymers; the Grashof and Rayleigh numbers Gr and Ra, describing density-driven flows; and the Knudsen number, describing the importance of noncontinuum molecular effects. Furthermore, the long-range nature of viscous flows and the small device dimensions inherent in microfluidics mean that the influence of boundaries is typically significant. A variety of strategies have been developed to manipulate fluids by exploiting boundary effects; among these are electrokinetic effects, acoustic streaming, and fluid-structure interactions. The goal is to describe the physics behind the rich variety of fluid phenomena occurring on the nanoliter scale using simple scaling arguments, with the hopes of developing an intuitive sense for this occasionally counterintuitive world.

Overstretching B-DNA: The Elastic Response of Individual Double-Stranded and Single-Stranded DNA Molecules

Abstract: Single molecules of double-stranded DNA (dsDNA) were stretched with force-measuring laser tweezers. Under a longitudinal stress of approximately 65 piconewtons (pN), dsDNA molecules in aqueous buffer undergo a highly cooperative transition into a stable form with 5.8 angstroms rise per base pair, that is, 70% longer than B form dsDNA. When the stress was relaxed below 65 pN, the molecules rapidly and reversibly contracted to their normal contour lengths. This transition was affected by changes in the ionic strength of the medium and the water activity or by cross-linking of the two strands of dsDNA. Individual molecules of single-stranded DNA were also stretched giving a persistence length of 7.5 angstroms and a stretch modulus of 800 pN. The overstretched form may play a significant role in the energetics of DNA recombination.

DNA in a material world

Abstract: The specific bonding of DNA base pairs provides the chemical foundation for genetics. This powerful molecular recognition system can be used in nanotechnology to direct the assembly of highly structured materials with specific nanoscale features, as well as in DNA computation to process complex information. The exploitation of DNA for material purposes presents a new chapter in the history of the molecule.

Single-molecule force spectroscopy: optical tweezers, magnetic tweezers and atomic force microscopy

Abstract: Single-molecule force spectroscopy has emerged as a powerful tool to investigate the forces and motions associated with biological molecules and enzymatic activity. The most common force spectroscopy techniques are optical tweezers, magnetic tweezers and atomic force microscopy. Here we describe these techniques and illustrate them with examples highlighting current capabilities and limitations.

A DNA-fuelled molecular machine made of DNA

Abstract: Molecular recognition between complementary strands of DNA allows construction on a nanometre length scale. For example, DNA tags may be used to organize the assembly of colloidal particles, and DNA templates can direct the growth of semiconductor nanocrystals and metal wires. As a structural material in its own right, DNA can be used to make ordered static arrays of tiles, linked rings and polyhedra. The construction of active devices is also possible--for example, a nanomechanical switch, whose conformation is changed by inducing a transition in the chirality of the DNA double helix. Melting of chemically modified DNA has been induced by optical absorption, and conformational changes caused by the binding of oligonucleotides or other small groups have been shown to change the enzymatic activity of ribozymes. Here we report the construction of a DNA machine in which the DNA is used not only as a structural material, but also as 'fuel'. The machine, made from three strands of DNA, has the form of a pair of tweezers. It may be closed and opened by addition of auxiliary strands of 'fuel' DNA; each cycle produces a duplex DNA waste product.