Functional expression and characterization of plant ABC transporters in Xenopus laevis oocytes for transport engineering purposes
Research output: Chapter in Book/Report/Conference proceeding › Book chapter › Research › peer-review
Standard
Functional expression and characterization of plant ABC transporters in Xenopus laevis oocytes for transport engineering purposes. / Xu, Deyang; Veres, Dorottya; Belew, Zeinu Mussa; Olsen, Carl Erik; Nour-Eldin, Hussam Hassan; Halkier, Barbara Ann.
Synthetic biology and metabolic engineering in plants and microbes : Part B: Plant metabolism. 2016. p. 207-224 (Methods in Enzymology, Vol. 576).Research output: Chapter in Book/Report/Conference proceeding › Book chapter › Research › peer-review
Harvard
APA
Vancouver
Author
Bibtex
}
RIS
TY - CHAP
T1 - Functional expression and characterization of plant ABC transporters in Xenopus laevis oocytes for transport engineering purposes
AU - Xu, Deyang
AU - Veres, Dorottya
AU - Belew, Zeinu Mussa
AU - Olsen, Carl Erik
AU - Nour-Eldin, Hussam Hassan
AU - Halkier, Barbara Ann
PY - 2016
Y1 - 2016
N2 - Transport engineering in bioengineering is aimed at efficient export of the final product to reduce toxicity and feedback inhibition and to increase yield. The ATP-binding cassette (ABC) transporters with their highly diverse substrate specificity and role in cellular efflux are potentially suitable in transport engineering approaches, although their size and high number of introns make them notoriously difficult to clone. Here, we report a novel in planta “exon engineering” strategy for cloning of full-length coding sequence of ABC transporters followed by methods for biochemical characterization of ABC exporters in Xenopus oocytes. Although the Xenopus oocyte expression system is particularly suitable for expression of membrane proteins and powerful in screening for novel transporter activity, only few examples of successful expression of ABC transporter has been reported. This raises the question whether the oocytes system is suitable to express and characterize ABC transporters. Thus we have selected AtABCG25, previously characterized in insect cells as the exporter of commercially valuable abscisic acid—as case study for optimizing of characterization in Xenopus oocytes. The tools provided will hopefully contribute to more successful transport engineering in synthetic biology.
AB - Transport engineering in bioengineering is aimed at efficient export of the final product to reduce toxicity and feedback inhibition and to increase yield. The ATP-binding cassette (ABC) transporters with their highly diverse substrate specificity and role in cellular efflux are potentially suitable in transport engineering approaches, although their size and high number of introns make them notoriously difficult to clone. Here, we report a novel in planta “exon engineering” strategy for cloning of full-length coding sequence of ABC transporters followed by methods for biochemical characterization of ABC exporters in Xenopus oocytes. Although the Xenopus oocyte expression system is particularly suitable for expression of membrane proteins and powerful in screening for novel transporter activity, only few examples of successful expression of ABC transporter has been reported. This raises the question whether the oocytes system is suitable to express and characterize ABC transporters. Thus we have selected AtABCG25, previously characterized in insect cells as the exporter of commercially valuable abscisic acid—as case study for optimizing of characterization in Xenopus oocytes. The tools provided will hopefully contribute to more successful transport engineering in synthetic biology.
U2 - 10.1016/bs.mie.2016.04.001
DO - 10.1016/bs.mie.2016.04.001
M3 - Book chapter
C2 - 27480688
SN - 978-0-12-804539-8
T3 - Methods in Enzymology
SP - 207
EP - 224
BT - Synthetic biology and metabolic engineering in plants and microbes
ER -
ID: 162573709