TY - JOUR
T1 - Functional characterization of the microtubule-binding and -destabilizing domains of CPAP and d-SAS-4
AU - Hsu, Wen Bin
AU - Hung, Liang Yi
AU - Tang, Chieh Ju C.
AU - Su, Chia Li
AU - Chang, Yulin
AU - Tang, Tang K.
N1 - Funding Information:
We are grateful for Drs. Ruey-Hwa Chen and David Tu for their helpful comments and discussion. We also thank Dan Chamberlin for the English editing help. This work was supported by a grant from the National Science Council and an institutional grant from Academia Sinica, Taipei, Taiwan.
PY - 2008/8/15
Y1 - 2008/8/15
N2 - We previously identified a novel centrosomal protein CPAP, which carries a 112-residue motif that is essential for microtubule destabilization. In this report, we define both the microtubule (MT) binding and destabilizing domains in human CPAP and analyze the mutations that affect its MT-destabilizing activity. Analysis of a series of CPAP truncated proteins showed that the MT-binding domain (MBD; residues 423-607) of CPAP is located next to its MT-destabilizing domain (MDD; residues 311-422). Site-specific mutagenesis revealed that the mutations that either disrupt the α-helical structure (Y341P, I346P, L348P, and triple-P) or alter the charge property (KR377EE) of the MDD significantly affect its MT-destabilizing ability. The activity for binding to a tubulin heterodimer was also significantly reduced in KR377EE mutant. Furthermore, we have analyzed the putative function of Drosophila d-SAS-4, a distant relative of human CPAP, which shares a conserved ∼ 20-aa sequence with the MDD of CPAP. Our results show that mutations in this conserved sequence also eliminate d-SAS-4′s MT-destabilizing activity, suggesting that d-SAS-4 and CPAP may play similar roles within cells.
AB - We previously identified a novel centrosomal protein CPAP, which carries a 112-residue motif that is essential for microtubule destabilization. In this report, we define both the microtubule (MT) binding and destabilizing domains in human CPAP and analyze the mutations that affect its MT-destabilizing activity. Analysis of a series of CPAP truncated proteins showed that the MT-binding domain (MBD; residues 423-607) of CPAP is located next to its MT-destabilizing domain (MDD; residues 311-422). Site-specific mutagenesis revealed that the mutations that either disrupt the α-helical structure (Y341P, I346P, L348P, and triple-P) or alter the charge property (KR377EE) of the MDD significantly affect its MT-destabilizing ability. The activity for binding to a tubulin heterodimer was also significantly reduced in KR377EE mutant. Furthermore, we have analyzed the putative function of Drosophila d-SAS-4, a distant relative of human CPAP, which shares a conserved ∼ 20-aa sequence with the MDD of CPAP. Our results show that mutations in this conserved sequence also eliminate d-SAS-4′s MT-destabilizing activity, suggesting that d-SAS-4 and CPAP may play similar roles within cells.
UR - https://www.scopus.com/pages/publications/48849104246
UR - https://www.scopus.com/pages/publications/48849104246#tab=citedBy
U2 - 10.1016/j.yexcr.2008.05.012
DO - 10.1016/j.yexcr.2008.05.012
M3 - Article
C2 - 18586240
AN - SCOPUS:48849104246
SN - 0014-4827
VL - 314
SP - 2591
EP - 2602
JO - Experimental Cell Research
JF - Experimental Cell Research
IS - 14
ER -