The anticancer natural product pironetin selectively targets Lys352 of alpha-tubulin.
Pironetin is a potent inhibitor of tubulin assembly and arrests cell cycle progression in M phase. Analyses of its structure-activity relationships suggested that pironetin covalently binds tubulin. To determine the binding site of pironetin, we synthesized biotinylated pironetin, which inhibited tubulin assembly both in vitro and in situ. The biotinylated pironetin selectively and covalently bound with tubulin. Partial digestion of biotinylated pironetin-treated tubulin by several proteases revealed that the binding site is the C-terminal portion of alpha-tubulin. By systematic alanine scanning, the pironetin binding site was determined to be Lys352 of alpha-tubulin. Lys352 is located at the entrance of a small pocket of alpha-tubulin, and this pocket faces the beta-tubulin of the next dimer. This is the first compound that covalently binds to the alpha subunit of tubulin and Lys352 of alpha-tubulin and inhibits the interaction of tubulin heterodimers.
Usui
Takeo
T
Antibiotics Laboratory, RIKEN Discovery Research Institute, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan.
Watanabe
Hiroyuki
H
Nakayama
Hiroshi
H
Tada
Yukio
Y
Kanoh
Naoki
N
Kondoh
Masuo
M
Asao
Tetsuji
T
Takio
Koji
K
Watanabe
Hidenori
H
Nishikawa
Kiyohiro
K
Kitahara
Takeshi
T
Osada
Hiroyuki
H
D000595:Amino Acid Sequence; D000818:Animals; D000970:Antineoplastic Agents / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D001665:Binding Sites / Q000187:drug effects; D002460:Cell Line; D002645:Chickens; D008239:Lysine / Q000737:chemistry*; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017434:Protein Structure, Tertiary; D011753:Pyrones / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D051381:Rats; D013329:Structure-Activity Relationship; D014404:Tubulin / Q000737:chemistry; D050257:Tubulin Modulators
D000970:Antineoplastic Agents; D011753:Pyrones; D014404:Tubulin; D050257:Tubulin Modulators; C088446:pironetin; D008239:Lysine