Characterization of two related Drosophila gamma-tubulin complexes that differ in their ability to nucleate microtubules.
gamma-tubulin exists in two related complexes in Drosophila embryo extracts (Moritz, M., Y. Zheng, B.M. Alberts, and K. Oegema. 1998. J. Cell Biol. 142:1- 12). Here, we report the purification and characterization of both complexes that we name gamma-tubulin small complex (gammaTuSC; approximately 280,000 D) and Drosophila gammaTuRC ( approximately 2,200,000 D). In addition to gamma-tubulin, the gammaTuSC contains Dgrip84 and Dgrip91, two proteins homologous to the Spc97/98p protein family. The gammaTuSC is a structural subunit of the gammaTuRC, a larger complex containing about six additional polypeptides. Like the gammaTuRC isolated from Xenopus egg extracts (Zheng, Y., M.L. Wong, B. Alberts, and T. Mitchison. 1995. Nature. 378:578-583), the Drosophila gammaTuRC can nucleate microtubules in vitro and has an open ring structure with a diameter of 25 nm. Cryo-electron microscopy reveals a modular structure with approximately 13 radially arranged structural repeats. The gammaTuSC also nucleates microtubules, but much less efficiently than the gammaTuRC, suggesting that assembly into a larger complex enhances nucleating activity. Analysis of the nucleotide content of the gammaTuSC reveals that gamma-tubulin binds preferentially to GDP over GTP, rendering gamma-tubulin an unusual member of the tubulin superfamily.
Oegema
K
K
Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, USA. Karen.Omega@EMBL-Heidelburg.DE
Wiese
C
C
Martin
O C
OC
Milligan
R A
RA
Iwamatsu
A
A
Mitchison
T J
TJ
Zheng
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D020285:Cryoelectron Microscopy; D004330:Drosophila / Q000235:genetics / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D019476:Insect Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D046911:Macromolecular Substances; D008870:Microtubules / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D014404:Tubulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014981:Xenopus
D019476:Insect Proteins; D046911:Macromolecular Substances; D014404:Tubulin; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate