Recombinant ALOD4 and OlyA Purification

ALOD4 construct was generously provided by the lab of Dr Arun Radhakrishnan. Recombinant His-tagged ALOD4 and OlyA were purified as previously described (16 (link)). Briefly, ALOD4 expression was induced with 1 mM IPTG in OD_0.5 BL21 (DE3) pLysS E. coli for 16 h at 18°C. Cells were lysed and His-ALOD4 and His-OlyA were isolated by nickel purification followed by size exclusion chromatography (HisTrap-HP Ni column, Tricorn 10/300 Superdex 200 gel filtration column; FPLC AKTA, GE Healthcare). Protein-rich fractions were pooled and concentration was measured using a NanoDrop instrument.

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Fowler J.W., Boutagy N.E., Zhang R., Horikami D., Whalen M.B., Romanoski C.E, & Sessa W.C. (2023). SREBP2 regulates the endothelial response to cytokines via direct transcriptional activation of KLF6. Journal of Lipid Research, 64(8), 100411.

Publication 2023

HisTrap-HP Ni column FPLC AKTA

Cells E coli Gel filtration Iptg Nickel Protein Size exclusion chromatography

Corresponding Organization : Yale University

Other organizations : University of Arizona

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Variable analysis

independent variables

IPTG concentration (1 mM)

dependent variables

ALOD4 and OlyA protein expression and purification

control variables

OD0.5 BL21 (DE3) pLysS E. coli cells
Temperature (18°C)
Incubation time (16 h)
Nickel purification and size exclusion chromatography

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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