Purification and NMR studies of RNA polymerase II C-terminal domain Phosphatase 1 containing ubiquitin like domain

Sunggeon Ko, Youngmin Lee, Jong Bok Yoon, Weontae Lee

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

RNA polymerase II C-terminal domain phosphatase 1 containing ubiquitin like domain (UBLCP1) has been identified as a regulatory molecule of RNA polymerase II. UBLCP1 consists of ubiquitin like domain (UBL) and phosphatase domain homologous with UDP and CTD phosphatase. UBLCP1 was cloned into the E.coli expression vectors, pET32a and pGEX 4T-1 with TEV protease cleavage site and purified using both affinity and gel-filtration chromatography. Domains of UBLCP1 protein were successfully purified as 7 mg/500 mL (UBLCP1, 36.78 KDa), 32 mg/500 mL (UBL, 9 KDa) and 8 mg/500 mL (phosphatase domain, 25 KDa) yielded in LB medium, respectively. Isotope-labeled samples including triple-labeled (2H/15N/13C) UBLCP1 were also prepared for hetero-nuclear NMR experiments. 15N-1H 2D-HSQC spectra of UBLCP1 suggest that both UBL and phosphatase domain are properly folded and structurally independent each other. These data will promise us further structural investigation of UBLCP1 by NMR spectroscopy and/or X-ray crystallography.

Original languageEnglish
Pages (from-to)1039-1042
Number of pages4
JournalBulletin of the Korean Chemical Society
Volume30
Issue number5
DOIs
Publication statusPublished - 2009 Oct 13

Fingerprint

RNA Polymerase II
Ubiquitin
Phosphoric Monoester Hydrolases
Purification
Nuclear magnetic resonance
Uridine Diphosphate
X ray crystallography
Chromatography
Isotopes
Escherichia coli
Nuclear magnetic resonance spectroscopy
Gels
Molecules
Proteins
Experiments

All Science Journal Classification (ASJC) codes

  • Chemistry(all)

Cite this

@article{124067c946f54cf5a813abb1612fb249,
title = "Purification and NMR studies of RNA polymerase II C-terminal domain Phosphatase 1 containing ubiquitin like domain",
abstract = "RNA polymerase II C-terminal domain phosphatase 1 containing ubiquitin like domain (UBLCP1) has been identified as a regulatory molecule of RNA polymerase II. UBLCP1 consists of ubiquitin like domain (UBL) and phosphatase domain homologous with UDP and CTD phosphatase. UBLCP1 was cloned into the E.coli expression vectors, pET32a and pGEX 4T-1 with TEV protease cleavage site and purified using both affinity and gel-filtration chromatography. Domains of UBLCP1 protein were successfully purified as 7 mg/500 mL (UBLCP1, 36.78 KDa), 32 mg/500 mL (UBL, 9 KDa) and 8 mg/500 mL (phosphatase domain, 25 KDa) yielded in LB medium, respectively. Isotope-labeled samples including triple-labeled (2H/15N/13C) UBLCP1 were also prepared for hetero-nuclear NMR experiments. 15N-1H 2D-HSQC spectra of UBLCP1 suggest that both UBL and phosphatase domain are properly folded and structurally independent each other. These data will promise us further structural investigation of UBLCP1 by NMR spectroscopy and/or X-ray crystallography.",
author = "Sunggeon Ko and Youngmin Lee and Yoon, {Jong Bok} and Weontae Lee",
year = "2009",
month = "10",
day = "13",
doi = "10.5012/bkcs.2009.30.5.1039",
language = "English",
volume = "30",
pages = "1039--1042",
journal = "Bulletin of the Korean Chemical Society",
issn = "0253-2964",
publisher = "Korean Chemical Society",
number = "5",

}

Purification and NMR studies of RNA polymerase II C-terminal domain Phosphatase 1 containing ubiquitin like domain. / Ko, Sunggeon; Lee, Youngmin; Yoon, Jong Bok; Lee, Weontae.

In: Bulletin of the Korean Chemical Society, Vol. 30, No. 5, 13.10.2009, p. 1039-1042.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Purification and NMR studies of RNA polymerase II C-terminal domain Phosphatase 1 containing ubiquitin like domain

AU - Ko, Sunggeon

AU - Lee, Youngmin

AU - Yoon, Jong Bok

AU - Lee, Weontae

PY - 2009/10/13

Y1 - 2009/10/13

N2 - RNA polymerase II C-terminal domain phosphatase 1 containing ubiquitin like domain (UBLCP1) has been identified as a regulatory molecule of RNA polymerase II. UBLCP1 consists of ubiquitin like domain (UBL) and phosphatase domain homologous with UDP and CTD phosphatase. UBLCP1 was cloned into the E.coli expression vectors, pET32a and pGEX 4T-1 with TEV protease cleavage site and purified using both affinity and gel-filtration chromatography. Domains of UBLCP1 protein were successfully purified as 7 mg/500 mL (UBLCP1, 36.78 KDa), 32 mg/500 mL (UBL, 9 KDa) and 8 mg/500 mL (phosphatase domain, 25 KDa) yielded in LB medium, respectively. Isotope-labeled samples including triple-labeled (2H/15N/13C) UBLCP1 were also prepared for hetero-nuclear NMR experiments. 15N-1H 2D-HSQC spectra of UBLCP1 suggest that both UBL and phosphatase domain are properly folded and structurally independent each other. These data will promise us further structural investigation of UBLCP1 by NMR spectroscopy and/or X-ray crystallography.

AB - RNA polymerase II C-terminal domain phosphatase 1 containing ubiquitin like domain (UBLCP1) has been identified as a regulatory molecule of RNA polymerase II. UBLCP1 consists of ubiquitin like domain (UBL) and phosphatase domain homologous with UDP and CTD phosphatase. UBLCP1 was cloned into the E.coli expression vectors, pET32a and pGEX 4T-1 with TEV protease cleavage site and purified using both affinity and gel-filtration chromatography. Domains of UBLCP1 protein were successfully purified as 7 mg/500 mL (UBLCP1, 36.78 KDa), 32 mg/500 mL (UBL, 9 KDa) and 8 mg/500 mL (phosphatase domain, 25 KDa) yielded in LB medium, respectively. Isotope-labeled samples including triple-labeled (2H/15N/13C) UBLCP1 were also prepared for hetero-nuclear NMR experiments. 15N-1H 2D-HSQC spectra of UBLCP1 suggest that both UBL and phosphatase domain are properly folded and structurally independent each other. These data will promise us further structural investigation of UBLCP1 by NMR spectroscopy and/or X-ray crystallography.

UR - http://www.scopus.com/inward/record.url?scp=70349734984&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=70349734984&partnerID=8YFLogxK

U2 - 10.5012/bkcs.2009.30.5.1039

DO - 10.5012/bkcs.2009.30.5.1039

M3 - Article

AN - SCOPUS:70349734984

VL - 30

SP - 1039

EP - 1042

JO - Bulletin of the Korean Chemical Society

JF - Bulletin of the Korean Chemical Society

SN - 0253-2964

IS - 5

ER -