(A) Domain organizations of ClpL, ClpG, ClpB. All AAA+ proteins consist of two AAA domains (AAA1, AAA2), a coiled-coil middle domain (M) and diverse N-terminal domains (N). ClpG additionally harbors …
The domain organization is indicated. Similar and identical residues are highlighted in light and dark blue.
(A) Relative luciferase disaggregation activities (% refolded luciferase/min) of ClpL in the presence of 2 mM ATP (and a regeneration mix), 2 mM ADP and without nucleotide were determined. The …
(A) Incubation of luciferase-YFP at 46°C only leads to unfolding of the luciferase moiety and the formation of mixed aggregates including folded YFP. Unfolding of YFP during disaggregation of …
Relative luciferase disaggregation activities (% refolded luciferase/min) of ClpL and L. monocytogenes (Lm) ClpB/KJE were determined. ClpL or DnaK were incubated for 30 min at the indicated …
(A) Disaggregation activities of ClpL and ΔN-ClpL toward aggregated luciferase and malate dehydrogenase (MDH) (each: % regain enzymatic activity/min) were determined. The activity of ClpL was set to …
(A) Domain organization of ClpL. Pore loop (Y170, Y504) and Walker B (E197, E530) residues of AAA1 and AAA2 domains are indicated. (B) ClpL ATPase activities were determined at 0.125 and 1 μM …
(A) E. coli dnaK103 cells harboring plasmids for IPTG-controlled expression of indicated disaggregases were grown at 30°C to mid-logarithmic growth phase and shifted to 49°C. Serial dilutions of …
Source data include non-cropped and non-processed images of SDS-gels and western blots.
Source data include non-cropped and non-processed images of SDS-gels and western blots and indicate sections and loading schemata of the respective figure supplement.
(A) AlphaFold2 model of ClpL NTD. The color code depicts the calculated confidence of the prediction (pLDDT). Residues that potentially participate in the formation of small hydrophobic cores (α1/2 …
(A) 1H,15N-HSQC of ClpL NTD with resonance assignment labeled at each peak. Several residues exhibit dispersed peak position, indicative of folding. (B) 1H,1H-2D NOESY in D2O of ClpL NTD confirming …
(A/D) ATPase activities of indicated ClpL mutants determined at 0.125 and 1 μM protein concentration. The ATPase activities of ClpL wild-type (WT) were set to 1. (B/C) Luciferase and malate …
(A) Varying the ratio of LN-ClpB* and ΔN-ClpB* leads to formation of mixed hexamers with diverse numbers of NTDs. (B) Luciferase disaggregation activities (% refolded luciferase/min) of mixed LN-ClpB…
(A) Relative luciferase disaggregation activities (% refolded luciferase/min) of indicated LN-ClpB* disaggregase mixtures were determined (DWB: ATPase-deficient LN-ClpB*-E279A/E678A, ΔN: ΔN-ClpB*). …
(A) AlphaFold2 model of L. monocytogenes (Lm) ClpL ring dimers. Positions of individual domains are indicated. (B) Negative stain EM of Lm ClpL. Two-dimensional (2D) class averages revealing single …
Source data includes the non-cropped and non-processed image of the SDS-gel.
Source data includes the non-cropped and non-processed image of the SDS-gel and indicates sections and loading schemata of the respective figure.
(A) Oligomeric states of ClpL wild-type (WT) and indicated mutants were determined in the presence of ATP by size exclusion chromatography. ATPase-deficient ClpB-E279A/E678A (DWB) served as …
Source data include non-cropped and non-processed images of SDS-gels.
Source data include non-cropped and non-processed images of SDS-gels and indicate sections and loading schemata of the respective figure supplement.
(A) Relative ATPase activities of indicated ClpL wild-type (WT) and T355C were determined at 0.125 and 1 μM protein concentration. The ATPase activities of ClpL-WT were set to 1. (B) Relative …
(A) Production levels of ClpL (wild-type [WT] and indicated mutants) in E. coli ΔclpB cells. E. coli ΔclpB cells harboring plasmids for IPTG-controlled expression of clpL (WT and indicated mutants) …
Source data includes the non-cropped and non-processed image of the SDS-gel.
Source data includes the non-cropped and non-processed image of the SDS-gel and indicates sections and loading schemata of the respective figure supplement.
coli dnaK103 mutant cells expressing E. coli dnaK, L. monocytogenes clpL or P. aeruginosa clpG were spotted on LB plates including the indicatedIPTG concentrations. Plates were incubated at 30°C or …
E. coli strains and plasmids used in this study.
Number of replicates.