title: ADP-ribosylation of platelet actin by botulinum C2 toxin.
abstract: Botulinum C2 toxin is a microbial toxin which possesses ADP-ribosyltransferase activity. In human platelet cytosol a 43-kDa protein was ADP-ribosylated by botulinum C2 toxin. Labelling of the 43-kDa protein using [32P]NAD as substrate was reduced by unlabelled NAD and nicotinamide. The label was removed by treatment with snake venom phosphodiesterase. Half-maximal and maximal ADP-ribosylation occurred at 0.1 microgram/ml and 3 micrograms/ml botulinum C2 toxin, respectively. The Km value of the ADP-ribosylation reaction for NAD was about 1 microM. The peptide map of the ADP-ribosylated 43-kDa protein was almost identical with platelet actin. The ADP-ribosylated 43-kDa substrate protein bound to and was eluted from immobilized DNase I in a manner similar to G-actin. Trypsin treatment of platelet cytosol decreased subsequent ADP-ribosylation of the 43-kDa protein without occurrence of smaller labelled polypeptides. Purified platelet actin was also ADP-ribosylated by botulinum C2 toxin with similar characteristics found with actin in platelet cytosol. Phalloidin decreased the ADP-ribosylation of actin in platelet cytosol and of isolated platelet actin. Half-maximal and maximal, about 90%, reduction of actin ADP-ribosylation was observed at 0.4 microM and 10 microM phalloidin, respectively. ADP-ribosylation of purified actin, induced by botulinum C2I toxin, abolished the formation of the typical microfilament network. The data indicate that platelet G-actin but not F-actin is a substrate of botulinum C2 toxin and that this covalent modification largely affects the functional properties of actin.
mesh_terms: D036002:ADP Ribose Transferases; D000199:Actins / Q000097:blood* / Q000502:physiology; D001792:Blood Platelets / Q000378:metabolism*; D001905:Botulinum Toxins / Q000494:pharmacology*; D002384:Catalysis; D003593:Cytoplasm / Q000378:metabolism; D003851:Deoxyribonucleases / Q000494:pharmacology; D008854:Microscopy, Electron; D009842:Oligopeptides / Q000494:pharmacology*; D010084:Oxidation-Reduction / Q000187:drug effects; D010430:Pentosyltransferases / Q000378:metabolism*; D010590:Phalloidine / Q000494:pharmacology*; D013379:Substrate Specificity; D014357:Trypsin / Q000494:pharmacology
chemical_list: D000199:Actins; D009842:Oligopeptides; D010590:Phalloidine; D036002:ADP Ribose Transferases; D010430:Pentosyltransferases; D003851:Deoxyribonucleases; D014357:Trypsin; D001905:Botulinum Toxins