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Evidence that the tightly bound magnesium in tubulin is associated with the N‐site GTP
Author(s) -
Osei Anthony A.,
Everett Grover W.,
Himes Richard H.
Publication year - 1990
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(90)80513-i
Subject(s) - gtp' , tubulin , chemistry , magnesium , nucleotide , phosphatase , alkaline phosphatase , biochemistry , microtubule , binding site , biophysics , enzyme , biology , microbiology and biotechnology , organic chemistry , gene
In an attempt to determine whether the tightly bound Mg 2+ 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 Mg 2+ content remained equal to about 1 tubulin under conditions where denaturation did not occur. Moreover, the Mg/GTP ratio always remained equal to 1. These results indicate that the Mg 2+ is associated with the N‐site GTP.