Insights into mechanism and functional consequences of heme binding to hemolysin-activating lysine acyltransferase HlyC from Escherichia coli

Publication date: September 2018
Source:Biochimica et Biophysica Acta (BBA) – General Subjects, Volume 1862, Issue 9
Author(s): Sandra Peherstorfer, Hans Henning Brewitz, Ajay Abisheck Paul George, Amelie Wißbrock, Jana Maria Adam, Lutz Schmitt, Diana Imhof
BackgroundTight regulation of heme homeostasis is a critical mechanism in pathogenic bacteria since heme functions as iron source and prosthetic group, but is also toxic at elevated concentrations. Hemolysin-activating lysine-acyltransferase (HlyC) from Escherichia coli is crucial for maturation of hemolysin A, which lyses several mammalian cells including erythrocytes liberating large amounts of heme for bacterial uptake. A possible impact and functional consequences of the released heme on events employing bacterial HlyC have remained unexplored.MethodsHeme binding to HlyC was investigated using UV/vis and SPR spectroscopy. Functional impact of heme association was examined using an in vitro hemolysis assay. The interaction was further studied by homology modeling, molecular docking and dynamics simulations.ResultsWe identified HlyC as potential heme-binding protein possessing heme-regulatory motifs. Using wild-type protein and a double alanine mutant we demonstrated that heme binds to HlyC via histidine 151 (H151). We could show further that heme inhibits the enzymatic activity of wild-type HlyC. Computational studies illustrated potential interaction sites in addition to H151 confirming the results from spectroscopy indicating more than one heme-binding site.ConclusionsTaken together, our results reveal novel insights into heme-protein interactions and regulation of a component of the heme uptake system in one of the major causative agents of urinary tract infections in humans.General significanceThis study points to a possible novel mechanism of regulation as present in many uropathogenic E. coli strains at an early stage of heme iron acquisition from erythrocytes for subsequent internalization by the bacterial heme-uptake machinery.

Graphical abstract

image