Purification and reconstitution of human membrane-bound DHRS7 (SDR34C1) from Sf9 cells

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216208%3A11160%2F14%3A10281983" target="_blank" >RIV/00216208:11160/14:10281983 - isvavai.cz</a>

Výsledek na webu

<a href="http://www.sciencedirect.com/science/article/pii/S104659281300257X" target="_blank" >http://www.sciencedirect.com/science/article/pii/S104659281300257X</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1016/j.pep.2013.11.013" target="_blank" >10.1016/j.pep.2013.11.013</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Purification and reconstitution of human membrane-bound DHRS7 (SDR34C1) from Sf9 cells

Popis výsledku v původním jazyce

Dehydrogenase/reductase SDR family member 7 (DHRS7, SDR34C1, retSDR4) is one of the many endoplasmic reticulum bound members of the SDR superfamily. Preliminary results indicate its potential significance in human metabolism. DHRS7 containing TEV-cleavable His(10) and FLAG-tag expressed in the Sf9 cell line was solubilised, purified, and reconstituted into liposomes to enable the improved characterisation of this enzyme in the future. Igepal CA-630 was determined to be the best detergent for the solubilisation process. The solubilised DHRS7 was purified using affinity chromatography, and the purified enzyme was subjected to TEV cleavage of the affinity tags and then repurified using subtractive Ni-IMAC. The cleaved and uncleaved versions of DHRS7 weresuccessfully reconstituted into liposomes. In addition, using tobacco specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) as the substrate, the cleaved liposomal DHRS7 was found to be inactive, whereas the pure and un

Název v anglickém jazyce

Purification and reconstitution of human membrane-bound DHRS7 (SDR34C1) from Sf9 cells

Popis výsledku anglicky

Dehydrogenase/reductase SDR family member 7 (DHRS7, SDR34C1, retSDR4) is one of the many endoplasmic reticulum bound members of the SDR superfamily. Preliminary results indicate its potential significance in human metabolism. DHRS7 containing TEV-cleavable His(10) and FLAG-tag expressed in the Sf9 cell line was solubilised, purified, and reconstituted into liposomes to enable the improved characterisation of this enzyme in the future. Igepal CA-630 was determined to be the best detergent for the solubilisation process. The solubilised DHRS7 was purified using affinity chromatography, and the purified enzyme was subjected to TEV cleavage of the affinity tags and then repurified using subtractive Ni-IMAC. The cleaved and uncleaved versions of DHRS7 weresuccessfully reconstituted into liposomes. In addition, using tobacco specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) as the substrate, the cleaved liposomal DHRS7 was found to be inactive, whereas the pure and un

Druh

J_x - Nezařazeno - Článek v odborném periodiku (Jimp, Jsc a Jost)

CEP obor

CE - Biochemie

OECD FORD obor

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Projekt

<a href="/cs/project/EE2.3.20.0235" target="_blank" >EE2.3.20.0235: Vybudování výzkumného týmu experimentální a aplikované biofarmacie</a><br>

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)<br>S - Specificky vyzkum na vysokych skolach<br>I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2014

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Protein Expression and Purification

ISSN

1046-5928

e-ISSN

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Svazek periodika

95

Číslo periodika v rámci svazku

March

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

6

Strana od-do

44-49

Kód UT WoS článku

000332192900007

EID výsledku v databázi Scopus

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