Evidence that the tightly bound magnesium in tubulin is associated with the N-site GTP.
In an attempt to determine whether the tightly bound Mg2+ found in purified tubulin in associated with the N-site GTP or the E-site GDP or GTP, we removed the E-site nucleotide by several means: (i) alkaline phosphatase treatment; (ii) displacement using excess GMPPCP; and (iii) polymerizing tubulin in the presence of alkaline phosphatase and non-hydrolyzable analogues. The Mg2+ content remained equal to about 1 mol/mol tubulin under conditions where denaturation did not occur. Moreover, the Mg/GTP ratio always remained equal to 1. These results indicate that the Mg2+ is associated with the N-site GTP.
Osei
A A
AA
Department of Biochemistry, University of Kansas, Lawrence 66045.
Everett
G W
GW
Himes
R H
RH
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D001921:Brain / Q000378:metabolism; D002417:Cattle; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D007700:Kinetics; D008274:Magnesium / Q000378:metabolism*; D014404:Tubulin / Q000378:metabolism*
D014404:Tubulin; D006160:Guanosine Triphosphate; D008274:Magnesium