[Fluorescent derivatives of diphtheria toxin subunit B and their interaction with Vero cells].
Diphtheria toxin's B subunit provides toxin interaction with its receptor on the cell surface and translocation of toxin's A subunit from endosome to cytozole of sensitive cells. Functional analogues of B subunit with fluorescent label are considered as perspective tools for studying the above mentioned processes. The aim of the work was to obtain fluorescent B subunit analogues and to detect the specificity of their interaction with Vero line cells. B subunit fluorescent analogues were obtained in two different ways. The first one was B subunit chemical conjugation with fluorescein isothiocyanate and the second one was genetic fusion of recombinant B subunit chain with enhanced green fluorescent protein chain. Specific interaction of B subunit fluorescent derivatives with Vero cells was studied by flow cytometry and confocal microscopy. Using competitive analysis it was shown that B subunit fluorescent analogues possessed different affinity for cells. The affinity of EGFP-SbB was higher than FITC-SbB. Our results indicate the possibility to use the fluorescent derivatives of B subunit as tools for identification of diphtheria toxin's receptor (HB-EGF) expression on the cell surface as well as for studying the interaction and penetration of diphtheria toxin to the cell.
Kaberniuk
A A
AA
Labyntsev
A Iu
AIu
Kolybo
D V
DV
Oliĭnyk
O S
OS
Redchuk
T A
TA
Korotkevych
N V
NV
Horchev
V F
VF
Karakhim
S O
SO
Komisarenko
S V
SV
D000818:Animals; D001667:Binding, Competitive; D018929:Cell Culture Techniques; D002522:Chlorocebus aethiops; D004167:Diphtheria Toxin / Q000235:genetics / Q000378:metabolism*; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D018613:Microscopy, Confocal; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D014709:Vero Cells
D004167:Diphtheria Toxin; D005456:Fluorescent Dyes; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C039171:diphtheria toxin fragment B; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D016650:Fluorescein-5-isothiocyanate