Showing papers by "Jens H. Gundlach published in 2015"
TL;DR: Ion current modulation through the protein nanopore MspA is used to observe translocation of helicase Hel308 on DNA with up to ∼40 pm sensitivity, applicable to any protein that translocates on DNA or RNA.
Abstract: Techniques for measuring the motion of single motor proteins, such as FRET and optical tweezers, are limited to a resolution of ∼300 pm. We use ion current modulation through the protein nanopore MspA to observe translocation of helicase Hel308 on DNA with up to ∼40 pm sensitivity. This approach should be applicable to any protein that translocates on DNA or RNA, including helicases, polymerases, recombinases and DNA repair enzymes.
108 citations
TL;DR: The dual mechanism of action of the bacterial pigment/lipid toxin leading to hemolysis or pyroptosis exacerbates fetal injury is demonstrated and it is suggested that preventing both activities of the hemolytic lipid is likely critical to reduce GBS fetal injury and preterm birth.
Abstract: Group B streptococci (GBS) are Gram-positive bacteria that cause infections in utero and in newborns. We recently showed that the GBS pigment is hemolytic and increased pigment production promotes bacterial penetration of human placenta. However, mechanisms utilized by the hemolytic pigment to induce host cell lysis and the consequence on fetal injury are not known. Here, we show that the GBS pigment induces membrane permeability in artificial lipid bilayers and host cells. Membrane defects induced by the GBS pigment trigger K + efflux leading to osmotic lysis of red blood cells or pyroptosis in human macrophages. Macrophages lacking the NLRP3 inflammasome recovered from pigment-induced cell damage. In a murine model of in utero infection, hyperpigmented GBS strains induced fetal injury in both an NLRP3 inflammasome-dependent and NLRP3 inflammasome-independent manner. These results demonstrate that the dual mechanism of action of the bacterial pigment/ lipid toxin leading to hemolysis or pyroptosis exacerbates fetal injury and suggest that preventing both activities of the hemolytic lipid is likely critical to reduce GBS fetal injury and preterm birth.
74 citations
TL;DR: In this article, the authors provide a comprehensive compilation of the results of these measurements, and also, using the dates of the measurements, consider whether $G$ is in fact a constant or if, instead, there is evidence for its variation with time.
Abstract: Newton's gravitational coupling, $G$, is a fundamental quantity that has been experimentally determined by many groups over the years. This paper not only provides a comprehensive compilation of the results of these measurements, but also, using the dates of the measurements, considers whether $G$ is in fact a constant or if, instead, there is evidence for its variation with time.
52 citations
TL;DR: It is found that both phi29 DNA polymerase and Hel308 helicase are capable of controlling the motion of DNA containing dNaM and d5SICS through the pore and that single reads are sufficient to detect the presence and location of dNaSICS within single molecules.
Abstract: Malyshev et al. showed that the four-letter genetic code within a living organism could be expanded to include the unnatural DNA bases dNaM and d5SICS. However, verification and detection of these unnatural bases in DNA requires new sequencing techniques. Here we provide proof of concept detection of dNaM and d5SICS in DNA oligomers via nanopore sequencing using the nanopore MspA. We find that both phi29 DNA polymerase and Hel308 helicase are capable of controlling the motion of DNA containing dNaM and d5SICS through the pore and that single reads are sufficient to detect the presence and location of dNaM and d5SICS within single molecules.
25 citations
TL;DR: It is demonstrated for the first time the effect of polymer composition on MspA protein functionality, and it is shown that membrane-protein interaction depends on the hydrophobic-hydrophilic ratio (f-ratio) of the block copolymer.
Abstract: Nanopores based on protein channels inserted into lipid membranes have paved the way towards a wide-range of inexpensive biosensors, especially for DNA sequencing. A key obstacle in using these biological ion channels as nanodevices is the poor stability of lipid bilayer membranes. Amphiphilic block copolymer membranes have emerged as a robust alternative to lipid membranes. While previous efforts have shown feasibility, we demonstrate for the first time the effect of polymer composition on MspA protein functionality. We show that membrane-protein interaction depends on the hydrophobic-hydrophilic ratio (f-ratio) of the block copolymer. These effects are particularly pronounced in asymmetric protein pores like MspA compared to the cylindrical α-Hemolysin pore. A key effect of membrane-protein interaction is the increased 1/fα noise. After first showing increases in 1/fα behaviour arise from increased substate activity, the noise power spectral density S(f) was used as a qualitative tool for understanding protein-membrane interactions in polymer membranes. Polymer compositions with f-ratios close to lipid membranes caused noise behaviour not observed in lipid membranes. However, by modifying the f-ratio using a modular synthetic approach, we were able to design a block copolymer exhibiting noise properties similar to a lipid membrane, albeit with better stability. Thus, by careful optimization, block copolymer membranes can emerge as a robust alternative for protein-pore based nano-biosensors.
23 citations