LIVIVO - The Search Portal for Life Sciences

zur deutschen Oberfläche wechseln
Advanced search

Search results

Result 1 - 6 of total 6

Search options

  1. Article ; Online: Single-protein detection in crowded molecular environments in cryo-EM images.

    Rickgauer, J Peter / Grigorieff, Nikolaus / Denk, Winfried

    eLife

    2017  Volume 6

    Abstract: We present an approach to study macromolecular assemblies by detecting component proteins' characteristic high-resolution projection patterns, calculated from their known 3D structures, in single electron cryo-micrographs. Our method detects single ... ...

    Abstract We present an approach to study macromolecular assemblies by detecting component proteins' characteristic high-resolution projection patterns, calculated from their known 3D structures, in single electron cryo-micrographs. Our method detects single apoferritin molecules in vitreous ice with high specificity and determines their orientation and location precisely. Simulations show that high spatial-frequency information and-in the presence of protein background-a whitening filter are essential for optimal detection, in particular for images taken far from focus. Experimentally, we could detect small viral RNA polymerase molecules, distributed randomly among binding locations, inside rotavirus particles. Based on the currently attainable image quality, we estimate a threshold for detection that is 150 kDa in ice and 300 kDa in 100 nm thick samples of dense biological material.
    Language English
    Publishing date 2017-05-03
    Publishing country England
    Document type Journal Article
    ZDB-ID 2687154-3
    ISSN 2050-084X ; 2050-084X
    ISSN (online) 2050-084X
    ISSN 2050-084X
    DOI 10.7554/eLife.25648
    Database MEDical Literature Analysis and Retrieval System OnLINE

    More links

    Kategorien

    Order via subito

    This service is chargeable due to the Delivery terms set by subito. Orders including an article and supplementary material will be classified as separate orders. In these cases, fees will be demanded for each order.

  2. Article ; Online: Single-protein detection in crowded molecular environments in cryo-EM images

    J Peter Rickgauer / Nikolaus Grigorieff / Winfried Denk

    eLife, Vol

    2017  Volume 6

    Abstract: We present an approach to study macromolecular assemblies by detecting component proteins’ characteristic high-resolution projection patterns, calculated from their known 3D structures, in single electron cryo-micrographs. Our method detects single ... ...

    Abstract We present an approach to study macromolecular assemblies by detecting component proteins’ characteristic high-resolution projection patterns, calculated from their known 3D structures, in single electron cryo-micrographs. Our method detects single apoferritin molecules in vitreous ice with high specificity and determines their orientation and location precisely. Simulations show that high spatial-frequency information and—in the presence of protein background—a whitening filter are essential for optimal detection, in particular for images taken far from focus. Experimentally, we could detect small viral RNA polymerase molecules, distributed randomly among binding locations, inside rotavirus particles. Based on the currently attainable image quality, we estimate a threshold for detection that is 150 kDa in ice and 300 kDa in 100 nm thick samples of dense biological material.
    Keywords optical methods ; cryo-EM ; Medicine ; R ; Science ; Q ; Biology (General) ; QH301-705.5
    Subject code 612
    Language English
    Publishing date 2017-05-01T00:00:00Z
    Publisher eLife Sciences Publications Ltd
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

    Full text online

    More links

    Kategorien

    Inter-library loan at ZB MED

    Your chosen title can be delivered directly to ZB MED Cologne location if you are registered as a user at ZB MED Cologne.

  3. Article ; Online: Portal motor velocity and internal force resisting viral DNA packaging in bacteriophage phi29.

    Rickgauer, John Peter / Fuller, Derek N / Grimes, Shelley / Jardine, Paul J / Anderson, Dwight L / Smith, Douglas E

    Biophysical journal

    2007  Volume 94, Issue 1, Page(s) 159–167

    Abstract: During the assembly of many viruses, a powerful molecular motor compacts the genome into a preassembled capsid. Here, we present measurements of viral DNA packaging in bacteriophage phi29 using an improved optical tweezers method that allows DNA ... ...

    Abstract During the assembly of many viruses, a powerful molecular motor compacts the genome into a preassembled capsid. Here, we present measurements of viral DNA packaging in bacteriophage phi29 using an improved optical tweezers method that allows DNA translocation to be measured from initiation to completion. This method allowed us to study the previously uncharacterized early stages of packaging and facilitated more accurate measurement of the length of DNA packaged. We measured the motor velocity versus load at near-zero filling and developed a ramped DNA stretching technique that allowed us to measure the velocity versus capsid filling at near-zero load. These measurements reveal that the motor can generate significantly higher velocities and forces than detected previously. Toward the end of packaging, the internal force resisting DNA confinement rises steeply, consistent with the trend predicted by many theoretical models. However, the force rises to a higher magnitude, particularly during the early stages of packaging, than predicted by models that assume coaxial inverse spooling of the DNA. This finding suggests that the DNA is not arranged in that conformation during the early stages of packaging and indicates that internal force is available to drive complete genome ejection in vitro. The maximum force exceeds 100 pN, which is about one-half that predicted to rupture the capsid shell.
    MeSH term(s) Bacillus Phages/physiology ; Computer Simulation ; DNA/physiology ; DNA Packaging/physiology ; Models, Biological ; Molecular Motor Proteins/physiology ; Motion ; Stress, Mechanical
    Chemical Substances Molecular Motor Proteins ; DNA (9007-49-2)
    Language English
    Publishing date 2007-09-07
    Publishing country United States
    Document type Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
    ZDB-ID 218078-9
    ISSN 1542-0086 ; 0006-3495
    ISSN (online) 1542-0086
    ISSN 0006-3495
    DOI 10.1529/biophysj.107.104612
    Database MEDical Literature Analysis and Retrieval System OnLINE

    More links

    Kategorien

    In stock of ZB MED Cologne/Königswinter

    Zs.A 235: Show issues Location:
    Je nach Verfügbarkeit (siehe Angabe bei Bestand)
    bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular
    Jg. 1995 - 2021: Lesesall (1.OG)
    ab Jg. 2022: Lesesaal (EG)
    Zs.MO 2: Show issues

    Order via subito

    This service is chargeable due to the Delivery terms set by subito. Orders including an article and supplementary material will be classified as separate orders. In these cases, fees will be demanded for each order.

  4. Article: Ionic effects on viral DNA packaging and portal motor function in bacteriophage phi 29.

    Fuller, Derek N / Rickgauer, John Peter / Jardine, Paul J / Grimes, Shelley / Anderson, Dwight L / Smith, Douglas E

    Proceedings of the National Academy of Sciences of the United States of America

    2007  Volume 104, Issue 27, Page(s) 11245–11250

    Abstract: In many viruses, DNA is confined at such high density that its bending rigidity and electrostatic self-repulsion present a strong energy barrier in viral assembly. Therefore, a powerful molecular motor is needed to package the DNA into the viral capsid. ... ...

    Abstract In many viruses, DNA is confined at such high density that its bending rigidity and electrostatic self-repulsion present a strong energy barrier in viral assembly. Therefore, a powerful molecular motor is needed to package the DNA into the viral capsid. Here, we investigate the role of electrostatic repulsion on single DNA packaging dynamics in bacteriophage phi 29 via optical tweezers measurements. We show that ionic screening strongly affects the packing forces, confirming the importance of electrostatic repulsion. Separately, we find that ions affect the motor function. We separate these effects through constant force measurements and velocity versus load measurements at both low and high capsid filling. Regarding motor function, we find that eliminating free Mg(2+) blocks initiation of packaging. In contrast, Na(+) is not required, but it increases the motor velocity by up to 50% at low load. Regarding internal resistance, we find that the internal force was lowest when Mg(2+) was the dominant ion or with the addition of 1 mM Co(3+). Forces resisting DNA confinement were up to approximately 80% higher with Na(+) as the dominant counterion, and only approximately 90% of the genome length could be packaged in this condition. The observed trend of the packing forces is in accord with that predicted by DNA charge-screening theory. However, the forces are up to six times higher than predicted by models that assume coaxial spooling of the DNA and interaction potentials derived from DNA condensation experiments. The forces are also severalfold higher than ejection forces measured with bacteriophage lambda.
    MeSH term(s) Bacillus Phages/chemistry ; Bacillus Phages/genetics ; Bacillus Phages/physiology ; Capsid/chemistry ; Capsid/physiology ; Cations, Divalent/chemistry ; Cations, Monovalent/chemistry ; Cobalt/chemistry ; Cobalt/physiology ; DNA, Viral/chemistry ; DNA, Viral/physiology ; Magnesium/chemistry ; Magnesium/physiology ; Optical Tweezers ; Predictive Value of Tests ; Sodium/chemistry ; Sodium/physiology ; Static Electricity ; Virus Assembly/physiology
    Chemical Substances Cations, Divalent ; Cations, Monovalent ; DNA, Viral ; Cobalt (3G0H8C9362) ; Sodium (9NEZ333N27) ; Magnesium (I38ZP9992A)
    Language English
    Publishing date 2007-06-07
    Publishing country United States
    Document type Comparative Study ; Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
    ZDB-ID 209104-5
    ISSN 1091-6490 ; 0027-8424
    ISSN (online) 1091-6490
    ISSN 0027-8424
    DOI 10.1073/pnas.0701323104
    Database MEDical Literature Analysis and Retrieval System OnLINE

    More links

    Kategorien

    In stock of ZB MED Cologne/Königswinter

    Zs.A 1148: Show issues Location:
    Je nach Verfügbarkeit (siehe Angabe bei Bestand)
    bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular
    Jg. 1995 - 2021: Lesesall (1.OG)
    ab Jg. 2022: Lesesaal (EG)

    Order via subito

    This service is chargeable due to the Delivery terms set by subito. Orders including an article and supplementary material will be classified as separate orders. In these cases, fees will be demanded for each order.

  5. Article ; Online: A general method for manipulating DNA sequences from any organism with optical tweezers.

    Fuller, Derek N / Gemmen, Gregory J / Rickgauer, John Peter / Dupont, Aurelie / Millin, Rachel / Recouvreux, Pierre / Smith, Douglas E

    Nucleic acids research

    2006  Volume 34, Issue 2, Page(s) e15

    Abstract: Mechanical manipulation of single DNA molecules can provide novel information about DNA properties and protein-DNA interactions. Here we describe and characterize a useful method for manipulating desired DNA sequences from any organism with optical ... ...

    Abstract Mechanical manipulation of single DNA molecules can provide novel information about DNA properties and protein-DNA interactions. Here we describe and characterize a useful method for manipulating desired DNA sequences from any organism with optical tweezers. Molecules are produced from either genomic or cloned DNA by PCR using labeled primers and are tethered between two optically trapped microspheres. We demonstrate that human, insect, plant, bacterial and viral sequences ranging from approximately 10 to 40 kilobasepairs can be manipulated. Force-extension measurements show that these constructs exhibit uniform elastic properties in accord with the expected contour lengths for the targeted sequences. Detailed protocols for preparing and manipulating these molecules are presented, and tethering efficiency is characterized as a function of DNA concentration, ionic strength and pH. Attachment strength is characterized by measuring the unbinding time as a function of applied force. An alternative stronger attachment method using an amino-carboxyl linkage, which allows for reliable DNA overstretching, is also described.
    MeSH term(s) Base Sequence ; DNA/chemistry ; DNA/isolation & purification ; Elasticity ; Genetic Techniques ; Humans ; Lasers ; Microfluidic Analytical Techniques ; Microspheres ; Polymerase Chain Reaction
    Chemical Substances DNA (9007-49-2)
    Language English
    Publishing date 2006-02-01
    Publishing country England
    Document type Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
    ZDB-ID 186809-3
    ISSN 1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
    ISSN (online) 1362-4962 ; 1362-4954
    ISSN 0301-5610 ; 0305-1048
    DOI 10.1093/nar/gnj016
    Database MEDical Literature Analysis and Retrieval System OnLINE

    More links

    Kategorien

    In stock of ZB MED Cologne/Königswinter

    Zs.A 1067: Show issues Location:
    Je nach Verfügbarkeit (siehe Angabe bei Bestand)
    bis Jg. 1994: Bestellungen von Artikeln über das Online-Bestellformular
    Jg. 1995 - 2021: Lesesall (1.OG)
    ab Jg. 2022: Lesesaal (EG)

    Order via subito

    This service is chargeable due to the Delivery terms set by subito. Orders including an article and supplementary material will be classified as separate orders. In these cases, fees will be demanded for each order.

  6. Article: Ionic effects on viral DNA packaging and portal motor function in bacteriophage φ29

    Fuller, Derek N / Rickgauer, John Peter / Jardine, Paul J / Grimes, Shelley / Anderson, Dwight L / Smith, Douglas E

    Proceedings of the National Academy of Sciences of the United States of America. 2007 July 3, v. 104, no. 27

    2007  

    Abstract: In many viruses, DNA is confined at such high density that its bending rigidity and electrostatic self-repulsion present a strong energy barrier in viral assembly. Therefore, a powerful molecular motor is needed to package the DNA into the viral capsid. ... ...

    Abstract In many viruses, DNA is confined at such high density that its bending rigidity and electrostatic self-repulsion present a strong energy barrier in viral assembly. Therefore, a powerful molecular motor is needed to package the DNA into the viral capsid. Here, we investigate the role of electrostatic repulsion on single DNA packaging dynamics in bacteriophage φ29 via optical tweezers measurements. We show that ionic screening strongly affects the packing forces, confirming the importance of electrostatic repulsion. Separately, we find that ions affect the motor function. We separate these effects through constant force measurements and velocity versus load measurements at both low and high capsid filling. Regarding motor function, we find that eliminating free Mg²⁺ blocks initiation of packaging. In contrast, Na⁺ is not required, but it increases the motor velocity by up to 50% at low load. Regarding internal resistance, we find that the internal force was lowest when Mg²⁺ was the dominant ion or with the addition of 1 mM Co³⁺. Forces resisting DNA confinement were up to [almost equal to]80% higher with Na⁺ as the dominant counterion, and only [almost equal to]90% of the genome length could be packaged in this condition. The observed trend of the packing forces is in accord with that predicted by DNA charge-screening theory. However, the forces are up to six times higher than predicted by models that assume coaxial spooling of the DNA and interaction potentials derived from DNA condensation experiments. The forces are also severalfold higher than ejection forces measured with bacteriophage λ.
    Language English
    Dates of publication 2007-0703
    Size p. 11245-11250.
    Publishing place National Academy of Sciences
    Document type Article
    ZDB-ID 209104-5
    ISSN 1091-6490 ; 0027-8424
    ISSN (online) 1091-6490
    ISSN 0027-8424
    Database NAL-Catalogue (AGRICOLA)

    More links

    Kategorien

    In stock of ZB MED Bonn / Germany

    Z 4' 63/44: Show issues
    87/25270: Show issues
    Qa 4' 169/3: Show issues
    Z 8.7: Show issues

    Order via subito

    This service is chargeable due to the Delivery terms set by subito. Orders including an article and supplementary material will be classified as separate orders. In these cases, fees will be demanded for each order.

    Inter-library loan at ZB MED

    Your chosen title can be delivered directly to ZB MED Cologne location if you are registered as a user at ZB MED Cologne.

To top