Purification, crystallization and initial crystallographic analysis of RNA polymerase holoenzyme from Thermus thermophilus

Marina N. Vassylyeva; Jookyung Lee; Shun Ichi Sekine; Oleg Laptenko; Seiki Kuramitsu; Takehiko Shibata; Yorinao Inoue; Sergei Borukhov; Dmitry G. Vassylyev; Shigeyuki Yokoyama

doi:10.1107/S0907444902011770

Purification, crystallization and initial crystallographic analysis of RNA polymerase holoenzyme from Thermus thermophilus

Marina N. Vassylyeva, Jookyung Lee, Shun Ichi Sekine, Oleg Laptenko, Seiki Kuramitsu, Takehiko Shibata, Yorinao Inoue, Sergei Borukhov, Dmitry G. Vassylyev, Shigeyuki Yokoyama

Research output: Contribution to journal › Article › peer-review

34 Scopus citations

Abstract

RNA polymerase holoenzyme from Thermus thermophilus, consisting of six protein subunits (α₂, β, β′, ω and σ⁷⁰) and having a total molecular mass of about 450 kDa, was purified and crystallized by the hanging-drop vapour-diffusion technique under mild near-physiological conditions. The crystals diffract beyond 3 Å resolution. Careful analysis of diffraction data revealed that the crystals belong to space group P3₂, with unit-cell parameters a = b = 236.35, c = 249.04 Å, and have perfect twinning along the threefold axis. A complete data set at 3 Å resolution was collected and an unambiguous molecular-replacement solution was found using the structure of T. aquaticus RNA polymerase core enzyme as a search model. The refinement of structure and model building of the σ⁷⁰ subunit is now in progress.

Original language	English (US)
Pages (from-to)	1497-1500
Number of pages	4
Journal	Acta Crystallographica Section D: Biological Crystallography
Volume	58
Issue number	9
DOIs	https://doi.org/10.1107/S0907444902011770
State	Published - Sep 2002
Externally published	Yes

All Science Journal Classification (ASJC) codes

Structural Biology

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10.1107/S0907444902011770

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Vassylyeva, M. N., Lee, J., Sekine, S. I., Laptenko, O., Kuramitsu, S., Shibata, T., Inoue, Y., Borukhov, S., Vassylyev, D. G., & Yokoyama, S. (2002). Purification, crystallization and initial crystallographic analysis of RNA polymerase holoenzyme from Thermus thermophilus. Acta Crystallographica Section D: Biological Crystallography, 58(9), 1497-1500. https://doi.org/10.1107/S0907444902011770

Vassylyeva, Marina N. ; Lee, Jookyung ; Sekine, Shun Ichi ; Laptenko, Oleg ; Kuramitsu, Seiki ; Shibata, Takehiko ; Inoue, Yorinao ; Borukhov, Sergei ; Vassylyev, Dmitry G. ; Yokoyama, Shigeyuki. / Purification, crystallization and initial crystallographic analysis of RNA polymerase holoenzyme from Thermus thermophilus. In: Acta Crystallographica Section D: Biological Crystallography. 2002 ; Vol. 58, No. 9. pp. 1497-1500.

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title = "Purification, crystallization and initial crystallographic analysis of RNA polymerase holoenzyme from Thermus thermophilus",

abstract = "RNA polymerase holoenzyme from Thermus thermophilus, consisting of six protein subunits (α2, β, β′, ω and σ70) and having a total molecular mass of about 450 kDa, was purified and crystallized by the hanging-drop vapour-diffusion technique under mild near-physiological conditions. The crystals diffract beyond 3 {\AA} resolution. Careful analysis of diffraction data revealed that the crystals belong to space group P32, with unit-cell parameters a = b = 236.35, c = 249.04 {\AA}, and have perfect twinning along the threefold axis. A complete data set at 3 {\AA} resolution was collected and an unambiguous molecular-replacement solution was found using the structure of T. aquaticus RNA polymerase core enzyme as a search model. The refinement of structure and model building of the σ70 subunit is now in progress.",

author = "Vassylyeva, {Marina N.} and Jookyung Lee and Sekine, {Shun Ichi} and Oleg Laptenko and Seiki Kuramitsu and Takehiko Shibata and Yorinao Inoue and Sergei Borukhov and Vassylyev, {Dmitry G.} and Shigeyuki Yokoyama",

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doi = "10.1107/S0907444902011770",

language = "English (US)",

volume = "58",

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Vassylyeva, MN, Lee, J, Sekine, SI, Laptenko, O, Kuramitsu, S, Shibata, T, Inoue, Y, Borukhov, S, Vassylyev, DG & Yokoyama, S 2002, 'Purification, crystallization and initial crystallographic analysis of RNA polymerase holoenzyme from Thermus thermophilus', Acta Crystallographica Section D: Biological Crystallography, vol. 58, no. 9, pp. 1497-1500. https://doi.org/10.1107/S0907444902011770

Purification, crystallization and initial crystallographic analysis of RNA polymerase holoenzyme from Thermus thermophilus. / Vassylyeva, Marina N.; Lee, Jookyung; Sekine, Shun Ichi; Laptenko, Oleg; Kuramitsu, Seiki; Shibata, Takehiko; Inoue, Yorinao; Borukhov, Sergei; Vassylyev, Dmitry G.; Yokoyama, Shigeyuki.

In: Acta Crystallographica Section D: Biological Crystallography, Vol. 58, No. 9, 09.2002, p. 1497-1500.

Research output: Contribution to journal › Article › peer-review

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AU - Lee, Jookyung

AU - Sekine, Shun Ichi

AU - Laptenko, Oleg

AU - Kuramitsu, Seiki

AU - Shibata, Takehiko

AU - Inoue, Yorinao

AU - Borukhov, Sergei

AU - Vassylyev, Dmitry G.

AU - Yokoyama, Shigeyuki

PY - 2002/9

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AB - RNA polymerase holoenzyme from Thermus thermophilus, consisting of six protein subunits (α2, β, β′, ω and σ70) and having a total molecular mass of about 450 kDa, was purified and crystallized by the hanging-drop vapour-diffusion technique under mild near-physiological conditions. The crystals diffract beyond 3 Å resolution. Careful analysis of diffraction data revealed that the crystals belong to space group P32, with unit-cell parameters a = b = 236.35, c = 249.04 Å, and have perfect twinning along the threefold axis. A complete data set at 3 Å resolution was collected and an unambiguous molecular-replacement solution was found using the structure of T. aquaticus RNA polymerase core enzyme as a search model. The refinement of structure and model building of the σ70 subunit is now in progress.

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Purification, crystallization and initial crystallographic analysis of RNA polymerase holoenzyme from Thermus thermophilus

Abstract

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