Journal: Bioinformatics

Article Title: Differential proteostatic regulation of insoluble and abundant proteins

doi: 10.1093/bioinformatics/btz214

Figure Lengend Snippet: Experimental analysis of codon usage in the chaperone dependence of GFP. ( A ) Decoding times of our GFP variants. Decoding time is shown per residue position, with each panel representing one of the variants. ( B ) Representative western blot of the solubility analysis of GFP variants upon cell-free expression. Bands show Total (T) and Soluble (S) fractions after centrifugation at 21 000 g for 30 min. ‘+ Dnak’ and ‘+ GroEL’ indicate addition of either DnaK mix—containing DnaK, DnaJ and GrpE—or GroE mix—containing GroEL and GroES—respectively. ( C ) Bar plots showing mean solubility as determined through quantification of western blot bands in (B), whiskers indicate standard deviation ( n = 5). Statistical significance was determined through two-way ANOVA followed by Tukey’s post-hoc test (‘**’ indicates P -value <0.0021). ( D ) Representative western blot of the solubility analysis of GFP variants upon overexpression in E.coli K12. Bands show Soluble (S) and Insoluble (I) fractions after centrifugation at 17 100 g for 15 min. ‘+ Dnak’ indicates co-expression of pKJE7, encoding DnaK, DnaJ and GrpE. ( E ) Bar plots showing mean solubility as determined through quantification of western blot bands in (B), whiskers indicate standard deviation ( n = 4). Statistical significance was determined through two-way ANOVA followed by Bonferroni’s post-hoc test (‘**’ indicates P -value < 0.0021, ‘****’ indicates P -value < 0.0001). ( F ) Structured Illumination Microscopy images of E.coli after 3 h of expression of one of the GFP variants. GFP fluorescence is show in green. ‘+ Dnak’ indicates co-expression of pKJE7, encoding DnaK, DnaJ and GrpE. White arrows indicate the position of non-fluorescent inclusion bodies. The inset in the top center panel shows an overlay of intrinsic GFP fluorescence in green, and ReASH-EDT2 labeled GFP fluorescence in red

Article Snippet: Structured Illumination Microscopy was performed at the VIB Bio-imaging Core at KU Leuven.

Techniques: Residue, Western Blot, Solubility, Expressing, Centrifugation, Standard Deviation, Over Expression, Microscopy, Fluorescence, Labeling