Nontypeable Haemophilus influenzae-binding gangliosides of human respiratory (HEp-2) cells have a requisite lacto/neolacto core structure.
Nontypeable Haemophilus influenzae (NTHI) are a major cause of human infections. We previously demonstrated high affinity and high specificity binding of NTHI to minor gangliosides of human respiratory (HEp-2) cells and macrophages, but not to brain gangliosides. We further identified the NTHI-binding ganglioside of human macrophages as alpha2,3-sialylosylparagloboside (IV3NeuAc-nLcOse4Cer, nLM1), which possesses a neolacto core structure that is absent in brain gangliosides. This supported a hypothesis that lacto/neolacto core carbohydrates are critical for NTHI-ganglioside binding. To investigate, we determined the core carbohydrate structure of NTHI-binding gangliosides of HEp-2 cells, through multiple approaches, including specific enzymatic degradation, mass spectral analysis and gas-liquid chromatography. Our analyses denote the following critical structural attributes of NTHI-binding gangliosides: (1) a conserved lacto/neolacto core structure; (2) requisite sialylation, which may be either internal or external, with alpha2,3 (human macrophages) or alpha2,6 (HEp-2 cells) anomeric linkages; (3) internalized galactose residues. Mass spectral and gas chromatographic analyses confirm that NTHI-binding gangliosides of HEp-2 cells possess lacto/neolacto carbohydrate cores and identify the structure of the major peak as NeuAcalpha2-6Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glcbeta1-1Cer (alpha2,6-sialosylparagloboside, nLM1). Collectively, our studies denote NTHI-binding gangliosides as lacto/neolacto series structures.
Berenson
Charles S
CS
Infectious Disease Division, Department of Veterans Affairs Western New York Healthcare System, State University of New York at Buffalo School of Medicine, Buffalo, New York 14215, USA. berenson@acsu.buffalo.edu
Sayles
Kelly B
KB
Huang
Jing
J
Reinhold
Vernon N
VN
Garlipp
Mary Alice
MA
Yohe
Herbert C
HC
D001422:Bacterial Adhesion; D002460:Cell Line; D002849:Chromatography, Gas; D003016:Clostridium perfringens / Q000201:enzymology; D005732:Gangliosides / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D006193:Haemophilus influenzae / Q000145:classification / Q000276:immunology* / Q000378:metabolism* / Q000472:pathogenicity; D006801:Humans; D013058:Mass Spectrometry; D015394:Molecular Structure; D009439:Neuraminidase; D009522:Newcastle disease virus / Q000201:enzymology; D012137:Respiratory System / Q000276:immunology / Q000382:microbiology; D013329:Structure-Activity Relationship; D001616:beta-Galactosidase
D005732:Gangliosides; C036208:LM1 ganglioside; D009439:Neuraminidase; D001616:beta-Galactosidase