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Updated: Aug 12, 2026

Comparing the Affinity of GTPase-binding Proteins using Competition Assays
Published on: October 8, 2015
Evidence that the tightly bound magnesium in tubulin is associated with the N-site GTP
A A Osei1, G W Everett, R H Himes
1Department of Biochemistry, University of Kansas, Lawrence 66045.
Abstract:
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.
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