RT Journal Article SR Electronic T1 Cevipabulin-tubulin complex reveals a novel agent binding site on α-tubulin and provides insights into microtubule dynamic instability JF bioRxiv FD Cold Spring Harbor Laboratory SP 2020.09.11.293563 DO 10.1101/2020.09.11.293563 A1 Jianhong Yang A1 Yamei Yu A1 Yong Li A1 Haoyu Ye A1 Wei Yan A1 Lu Niu A1 Yunhua Zheng A1 Zhoufeng Wang A1 Zhuang Yang A1 Heying Pei A1 Haoche Wei A1 Min Zhao A1 Jiaolin Wen A1 Linyu Yang A1 Liang Ouyang A1 Yuquan Wei A1 Qiang Chen A1 Weimin Li A1 Lijuan Chen YR 2020 UL http://biorxiv.org/content/early/2020/09/11/2020.09.11.293563.abstract AB Microtubule, composed of αβ-tubulin heterodimers, remains as one of the most popular anticancer targets for decades. To date, anti-microtubule drugs mainly target β-tubulin to inhibit microtubule dynamic instability (MDI) while agents binding to α-tubulin are less well characterized and also the molecular mechanism of MDI is far from being articulated. Cevipabulin, an oral microtubule-active antitumor clinical candidate, is widely accepted as a microtubule stabilizing agent (MSA) but binds to the microtubule-destabilization vinblastine site on β-tubulin and this unusual phenomenon has so far failed to be explained. Our X-ray crystallography study reveals that, in addition binding to the vinblastine site, cevipabulin also binds to a novel site on α-tubulin (named the seventh site) which located at the region spatially corresponding to the vinblastine site on β-tubulin. Interestingly, cevipabulin exhibits two unique site-dependent functions. Cevipabulin binding to the seventh site promotes tubulin degradation through interaction of the non-exchengeable GTP to reduce tubulin stability. Cevipabulin binding to the vinblastine site enhances longitudinal interactions but inhibits lateral interactions of tubulins, thus inducing tubulin protofilament polymerization (but not microtubule polymerization like MSAs), and then tangling into irregular tubulin aggregates. Importantly, the tubulin-cevipabulin structure is an intermediate between “bent” and “straight” tubulins and the involved bent-to-straight conformation change will be helpful to fully understand the molecular mechanism of tubulin assembly. Our findings confirm cevipabulin is not an MSA and shed light on the development of a new generation of anti-microtubule drugs targeting the novel site on α-tubulin and also provide new insights into MDI.Competing Interest StatementThe authors have declared no competing interest.