title: Chelerythrine inhibits the sarco/endoplasmic reticulum Ca(2+)-ATPase and results in cell Ca(2+) imbalance.
abstract: The isoquinoline alkaloid chelerythrine is described as an inhibitor of SERCA. The ATPase inhibition presented two non-competitive components, Ki1=1, 2 μM and Ki2=26 μM. Conversely, chelerythrine presented a dual effect on the p-nitrophenylphosphatase (pNPPase) of SERCA. Ca(2+)-dependent pNPPase was activated up to ∼5 μM chelerythrine with inhibition thereafter. Ca(2+)-independent pNPPase was solely inhibited. The phosphorylation of SERCA with ATP reached half-inhibition with 10 μM chelerythrine and did not parallel the decrease of ATPase activity. In contrast, chelerythrine up to 50 μM increased the phosphorylation by Pi. Cross-linking of SERCA with glutaraldehyde was counteracted by high concentrations of chelerythrine. The controlled tryptic digestion of SERCA shows that the low-affinity binding of chelerythrine evoked an E2-like pattern. Our data indicate a non-competitive inhibition of ATP hydrolysis that favors buildup of the E2-conformers of the enzyme. Chelerythrine as low as 0.5-1.5 μM resulted in an increase of intracellular Ca(2+) on cultured PBMC cells. The inhibition of SERCA and the loss of cell Ca(2+) homeostasis could in part be responsible for some described cytotoxic effects of the alkaloid. Thus, the choice of chelerythrine as a PKC-inhibitor should consider its potential cytotoxicity due to the alkaloid's effects on SERCA.
authors.affiliation: Instituto de Bioquímica Médica Leopoldo de Meis, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; Laboratório de Toxinologia, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brazil. Electronic address: saulomvieira@gmail.com.
authors.forename: Saulo Martins
authors.initials: SM
authors.lastname: Vieira
authors.affiliation: Instituto de Bioquímica Médica Leopoldo de Meis, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
authors.forename: Vanessa Honorato
authors.initials: VH
authors.lastname: de Oliveira
authors.affiliation: Instituto de Bioquímica Médica Leopoldo de Meis, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; Instituto de Biofísica Carlos Chagas Filho, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.
authors.forename: Raphael do Carmo
authors.initials: Rdo C
authors.lastname: Valente
authors.affiliation: Laboratório de Biologia Molecular e Doenças Endêmicas, Instituto Oswaldo Cruz, FIOCRUZ, Rio de Janeiro, Brazil.
authors.forename: Otacílio da Cruz
authors.initials: Oda C
authors.lastname: Moreira
authors.affiliation: Instituto de Bioquímica Médica Leopoldo de Meis, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: cfontes@bioqmed.ufrj.br.
authors.forename: Carlos Frederico Leite
authors.initials: CF
authors.lastname: Fontes
authors.affiliation: Instituto de Bioquímica Médica Leopoldo de Meis, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil. Electronic address: jmignaco@bioqmed.ufrj.br.
authors.forename: Julio Alberto
authors.initials: JA
authors.lastname: Mignaco
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry; D000818:Animals; D053119:Benzophenanthridines / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D005976:Glutaral / Q000737:chemistry; D006801:Humans; D006868:Hydrolysis; D020128:Inhibitory Concentration 50; D007963:Leukocytes, Mononuclear / Q000166:cytology; D009000:Monocytes / Q000378:metabolism; D018482:Muscle, Skeletal / Q000201:enzymology; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000378:metabolism*; D014357:Trypsin / Q000737:chemistry
chemical_list: D053119:Benzophenanthridines; D000255:Adenosine Triphosphate; C016299:chelerythrine; D014357:Trypsin; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D002118:Calcium; D005976:Glutaral
keywords: ATPase; Calcium; Chelerythrine; Cytotoxicity; PBMC; SERCA