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. 2021 Dec 23;12:781857. doi: 10.3389/fpls.2021.781857

Figure 3.

Figure 3

Sucrose gradient interaction assay of Alb3 variants and Alb4 reconstituted in nanodiscs with chloroplast ribosomes. (A) Schematic representation of recombinant His-tagged full-length mature Alb3 (amino acids 55–462), Alb3ΔC (amino acids 55–369) and Alb4 (amino acids 46–499) reconstituted in MSP1D1 nanodiscs (NDs; not to scale) with asolectin as bilayer forming lipid. (B) Nanodiscs (NDs) containing Alb3 (Alb3-NDs), Alb4 (Alb4-NDs), or Alb3ΔC (Alb3ΔC-NDs) and NDs without inserted membrane protein as negative control were incubated with chloroplast ribosomes and loaded onto a sucrose density gradient. After ultracentrifugation, the gradient fractions were analyzed immunologically using antibodies against the His-tag (α-His), A. thaliana chloroplast Alb3 (α-Alb3), human Apolipoprotein A1 (α-ApoA1; the wildtype template of MSP1D1) and the chloroplast ribosomal protein uL4c (α-uL4c). Sucrose density gradient centrifugation of the Alb constructs, containing NDs in the absence of ribosomes was used as negative control.