Molecular architecture of the Jumonji C family histone demethylase KDM5B

Abstract The full length human histone 3 lysine 4 demethylase KDM5B (PLU-1/Jarid1B) has been studied using Hydrogen/Deuterium exchange mass spectrometry, homology modelling, sequence analysis, small angle X-ray scattering and electron microscopy. This first structure on an intact multi-domain Jumonji histone demethylase reveal that the so-called PLU region, in the central region of KDM5B, has a curved α-helical three-dimensional structure, that acts as a rigid linker between the catalytic core and a region comprising four α-helices, a loop comprising the PHD2 domain, two large intrinsically disordered loops and the PHD3 domain in close proximity. The dumbbell shaped and curved KDM5B architecture observed by electron microscopy is complementary to the nucleosome surface and has a striking overall similarity to that of the functionally related KDM1A/CoREST complex. This could suggest that there are similarities between the demethylation mechanisms employed by the two histone 3 lysine 4 demethylases at the molecular level

Salt-bridge Swapping in the EXXERFXYY Motif of Proton Coupled Oligopeptide Transporters

Proton-coupled oligopeptide transporters (POTs) couple the inward transport of di- or tripeptides with an inwardly directed transport of protons. Evidence from several studies of different POTs has pointed toward involvement of a highly conserved sequence motif, E(1)XXE(2)RFXYY (from here on referred to as E(1)XXE(2)R), located on Helix I, in interactions with the proton. In this study, we investigated the intracellular substrate accumulation by motif variants with all possible combinations of glutamate residues changed to glutamine and arginine changed to a tyrosine, the latter being a natural variant found in the Escherichia coli POT YjdL. We found that YjdL motif variants with E(1)XXE(2)R, E(1)XXE(2)Y, E(1)XXQ(2)Y, or Q(1)XXE(2)Y were able to accumulate peptide, whereas those with E(1)XXQ(2)R, Q(1)XXE(2)R, or Q(1)XXQ(2)Y were unable to accumulate peptide, and Q(1)XXQ(2)R abolished uptake. These results suggest a mechanism that involves swapping of an intramotif salt bridge, i.e. R-E(2) to R-E(1), which is consistent with previous structural studies. Molecular dynamics simulations of the motif variants E(1)XXE(2)R and E(1)XXQ(2)R support this mechanism. The simulations showed that upon changing conformation arginine pushes Helix V, through interactions with the highly conserved FYING motif, further away from the central cavity in what could be a stabilization of an inward facing conformation. As E(2) has been suggested to be the primary site for protonation, these novel findings show how protonation may drive conformational changes through interactions of two highly conserved motifs