Estimating the Size of PfGCN5 Complex

To estimate the size of the PfGCN5 complex, nuclear extract from the PfGCN5::PTP was incubated with IgG beads, eluted by TEV protease cleavage as described above, and applied to a Superose 6 gel filtration column (GE Healthcare). Molecular mass standards (Gel Filtration Calibration Kit HMW, GE Healthcare) were run under the same conditions to estimate the size of the complex. The fractions were analyzed by Western blotting using antibodies against the PTP tag, PfPHD1 and PfPHD2, while HAT activity in the fractions was measured as described previously [57 (link)].

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Miao J., Wang C., Lucky A.B., Liang X., Min H., Adapa S.R., Jiang R., Kim K, & Cui L. (2021). A unique GCN5 histone acetyltransferase complex controls erythrocyte invasion and virulence in the malaria parasite Plasmodium falciparum. PLoS Pathogens, 17(8), e1009351.

Publication 2021

Superose 6 gel filtration column Gel Filtration Calibration Kit HMW

Antibodies Cleavage Gel filtration Tev protease

Corresponding Organization : University of South Florida

Other organizations : Center for Global Health

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Variable analysis

independent variables

Incubation of nuclear extract from PfGCN5::PTP with IgG beads

dependent variables

Size of the PfGCN5 complex estimated by Superose 6 gel filtration column
Presence of PTP tag, PfPHD1 and PfPHD2 proteins in the fractions analyzed by Western blotting
HAT activity in the fractions

control variables

Molecular mass standards (Gel Filtration Calibration Kit HMW, GE Healthcare) run under the same conditions

controls

Positive control: Molecular mass standards used to estimate the size of the PfGCN5 complex
Negative control: Not explicitly mentioned

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