Zampanolide, a potent new microtubule-stabilizing agent, covalently reacts with the taxane luminal site in tubulin α,β-heterodimers and microtubules.
Zampanolide and its less active analog dactylolide compete with paclitaxel for binding to microtubules and represent a new class of microtubule-stabilizing agent (MSA). Mass spectrometry demonstrated that the mechanism of action of both compounds involved covalent binding to β-tubulin at residues N228 and H229 in the taxane site of the microtubule. Alkylation of N228 and H229 was also detected in α,β-tubulin dimers. However, unlike cyclostreptin, the other known MSA that alkylates β-tubulin, zampanolide was a strong MSA. Modeling the structure of the adducts, using the NMR-derived dactylolide conformation, indicated that the stabilizing activity of zampanolide is likely due to interactions with the M-loop. Our results strongly support the existence of the luminal taxane site of microtubules in tubulin dimers and suggest that microtubule nucleation induction by MSAs may proceed through an allosteric mechanism.
School of Biological Sciences, Victoria University of Wellington, Wellington 6012, New Zealand.
Jessica J
JJ
Field
Benet
B
Pera
Enrique
E
Calvo
Angeles
A
Canales
Didier
D
Zurwerra
Chiara
C
Trigili
Javier
J
Rodríguez-Salarichs
Ruth
R
Matesanz
Arun
A
Kanakkanthara
St John
SJ
Wakefield
A Jonathan
AJ
Singh
Jesús
J
Jiménez-Barbero
Peter
P
Northcote
John H
JH
Miller
Juan Antonio
JA
López
Ernest
E
Hamel
Isabel
I
Barasoain
Karl-Heinz
KH
Altmann
José Fernando
JF
Díaz
D000970:Antineoplastic Agents / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D001665:Binding Sites / Q000187:drug effects; D001952:Bridged-Ring Compounds / Q000737:chemistry / Q000378:metabolism*; D049109:Cell Proliferation / Q000187:drug effects; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004354:Drug Screening Assays, Antitumor; D006801:Humans; D007700:Kinetics; D018942:Macrolides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D009682:Magnetic Resonance Spectroscopy; D008870:Microtubules / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D008958:Models, Molecular; D015394:Molecular Structure; D013329:Structure-Activity Relationship; D043823:Taxoids / Q000737:chemistry / Q000378:metabolism*; D014404:Tubulin / Q000737:chemistry* / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D001952:Bridged-Ring Compounds; D018942:Macrolides; D043823:Taxoids; D014404:Tubulin; C444900:zampanolide; C080625:taxane