Hydrolysis-deficient mosaic microtubules as faithful mimics of the GTP cap

被引:1
作者
Estevez-Gallego, Juan [1 ,2 ]
Blum, Thorsten B. [2 ]
Ruhnow, Felix [1 ]
Gili, Maria [1 ]
Speroni, Silvia [1 ]
Garcia-Castellanos, Raquel [1 ]
Steinmetz, Michel O. [2 ,3 ]
Surrey, Thomas [1 ,4 ,5 ]
机构
[1] Barcelona Inst Sci & Technol, Ctr Genom Regulat CRG, Barcelona, Spain
[2] Paul Scherrer Inst, Div Biol & Chem, Lab Biomol Res, Villigen, Switzerland
[3] Univ Basel, Biozentrum, Basel, Switzerland
[4] Univ Pompeu Fabra UPF, Barcelona, Spain
[5] ICREA, Pg Lluis Companys 23, Barcelona, Spain
基金
瑞士国家科学基金会; 欧洲研究理事会;
关键词
SLOWLY HYDROLYZABLE ANALOG; PLUS-END TRACKING; DYNAMIC INSTABILITY; BETA-TUBULIN; IN-VITRO; SIZE; POLYMERIZATION; MICROSCOPY; INFORMATION; TRANSITIONS;
D O I
10.1038/s41467-025-57555-6
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
A critical feature of microtubules is their GTP cap, a stabilizing GTP-tubulin rich region at growing microtubule ends. Microtubules polymerized in the presence of GTP analogs or from GTP hydrolysis-deficient tubulin mutants have been used as GTP-cap mimics for structural and biochemical studies. However, these analogs and mutants generate microtubules with diverse biochemical properties and lattice structures, leaving it unclear what is the most faithful GTP mimic and hence the structure of the GTP cap. Here, we generate a hydrolysis-deficient human tubulin mutant, alpha E254Q, with the smallest possible modification. We show that alpha E254Q-microtubules are stable, but still exhibit mild mutation-induced growth abnormalities. However, mixing two GTP hydrolysis-deficient tubulin mutants, alpha E254Q and alpha E254N, at an optimized ratio eliminates growth and lattice abnormalities, indicating that these 'mosaic microtubules' are faithful GTP cap mimics. Their cryo-electron microscopy structure reveals that longitudinal lattice expansion, but not protofilament twist, is the primary structural feature distinguishing the GTP-tubulin containing cap from the GDP-tubulin containing microtubule shaft. However, alterations in protofilament twist may be transiently needed to allow lattice compaction and GTP hydrolysis. Together, our results provide insights into the structural origin of GTP cap stability, the pathway of GTP hydrolysis and hence microtubule dynamic instability.
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页数:17
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