Fig. 2: NMR features of NDM-1 degradation in E coli.

a 2D ¹H-¹⁵N SOFAST HMQC spectra of apoNDM-1 in E. coli cells when Prc and DegP were overexpressed (black) and at endogenous protease levels (light brown). Cyan circles identify NMR resonances with chemical shift displacements characteristic of newly generated C-terminal sites. Black arrows denote cross-peaks from amide groups of disordered peptides. The dotted square indicates tryptophan HN indole signals from newly generated apoNDM-1 peptide fragments. The vertical cyan line indicates the artifactual set of signals (T1 noise) caused by the sharp and intense resonance of unlabeled DPA. b Schematic representation of the resulting chemical groups upon amide peptide bond cleavage. c, d Representative time course of periplasmic apoNDM-1 degradation followed by 1D ¹H NMR analysis of tryptophan HN indole signals (c). Experiment was performed on two independent bacterial samples. Tryptophan HN indole NMR signal amplitude plateaus at c.a. 6.5 h (d). Each value corresponds to the normalized intensity at each timepoint. Error bars represent the experimental noise in each spectrum. e ¹³C detected CBCACO (black) and CACO (cyan) NMR spectra of E. coli cells overexpressing NDM-1 and Prc/DegP after DPA treatment or in its absence (yellow). ¹³CO NMR signals above 180 ppm belong to newly generated C-terminal backbone carboxylates. Cα and Cβ chemical shifts in the second spectral dimension identify the type of amino acid side chain. (*) indicate unassigned minor sites. In all cases, NMR spectra were registered at 20 °C.