Ganglioside-binding specificities of E. coli enterotoxin LT-IIc: Importance of long-chain fatty acyl ceramide.
Bacterial heat-labile (LT) enterotoxins signal through tightly regulated interactions with host cell gangliosides. LT-IIa and LT-IIb of Escherichia coli bind preferentially to gangliosides with a NeuAcα2-3Galβ1-3GalNAc terminus, with key distinctions in specificity. LT-IIc, a newly discovered E. coli LT, is comprised of an A polypeptide with high homology, and a B polypeptide with moderate homology, to LT-IIa and LT-IIb. LT-IIc is less cytotoxic than LT-IIa and LT-IIb. We theorized that LT-IIc-host cell interaction is regulated by specific structural attributes of immune cell ganglioside receptors and designed experiments to test this hypothesis. Overlay immunoblotting to a diverse array of neural and macrophage gangliosides indicated that LT-IIc bound to a restrictive range of gangliosides, each possessing a NeuAcα2-3Galβ1-3GalNAc with a requisite terminal sialic acid. LT-IIc did not bind to GM1a with short-chain fatty acyl ceramides. Affinity overlay immunoblots, constructed to a diverse array of known ganglioside structures of murine peritoneal macrophages, established that LT-IIc bound to GM1a comprised of long-chain fatty acyl ceramides. Findings were confirmed with LT-IIc also binding to GM1a of RAW264.7 cells, comprised of a long-chain fatty acyl ceramide. Thus, LT-IIc-ganglioside binding differs distinctly from that of LT-IIa and LT-IIb. LT-IIc binding is not just dependent on carbohydrate composition, but also upon the orientation of the oligosaccharide portion of GM1a by the ceramide moiety. These studies are the first demonstration of LT-ganglioside dependence upon ceramide composition and underscore the contribution of long-chain fatty acyl ceramides to host cell interactions.
Division of Infectious Disease (151), Department of Veterans Affairs Western New York Healthcare System, State University of New York at Buffalo School of Medicine, 3495 Bailey Avenue, Buffalo, NY 14215, USA. berenson@buffalo.edu
Charles S
CS
Berenson
Hesham F
HF
Nawar
Ragina L
RL
Kruzel
Lorrie M
LM
Mandell
Terry D
TD
Connell
D000276:Adjuvants, Immunologic / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D001427:Bacterial Toxins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D002240:Carbohydrate Sequence; D002478:Cells, Cultured; D002518:Ceramides / Q000737:chemistry / Q000378:metabolism*; D004768:Enterotoxins / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D005732:Gangliosides / Q000737:chemistry / Q000378:metabolism*; D008264:Macrophages / Q000378:metabolism; D051379:Mice; D013379:Substrate Specificity
D000276:Adjuvants, Immunologic; D001427:Bacterial Toxins; D002518:Ceramides; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D005732:Gangliosides; C027538:heat-labile enterotoxin, E coli