D. Gingras et R. Beliveau, CYTOSOLIC PROTEINS OF 21-23 KDA ARE METHYLATED BY KIDNEY CORTEX MEMBRANE-ASSOCIATED C-TERMINAL CARBOXYL METHYLTRANSFERASES, Biochimica et biophysica acta. Protein structure and molecular enzymology, 1204(2), 1994, pp. 149-156
We have studied the effect of a soluble fraction from kidney cortex on
the C-terminal carboxyl methylation of 21-23 kDa proteins catalyzed b
y membrane-associated methyltransferases. Addition of soluble proteins
to isolated luminal, antiluminal and intracellular membranes resulted
in a large increase in the methylation of the membrane-associated 21-
23 kDa substrates. Fractionation of the soluble extract from the corte
x by Q-Sepharose anion exchange chromatography showed the presence of
two distinct peaks of proteins presenting stimulating activities, elut
ing at 0.15 M (peak I) and 0.4 M (peak II) NaCl, respectively. Both pe
aks eluted as proteins of apparent molecular sizes of 40 kDa upon Supe
rose 6 gel-filtration chromatography. No methylation activity towards
N-acetyl-S-trans, trans-farnesylcysteine (AFC), a good substrate for C
-terminal carboxyl methyltransferases, was associated with either peak
s. In contrast, the increase in methylation induced by these proteins
was strongly inhibited by AFC, suggesting that the methylation induced
by these factors occurred on C-terminal isoprenylated cysteine residu
es. Both partially purified proteins competitively inhibited the methy
lation of AFC by the membrane-associated enzymes; suggesting that they
may represent substrates for the methyltransferases. Immunoblotting o
f these partially purified soluble substrates with a rabbit polyclonal
antibody directed against the small G-protein CDC42 showed the presen
ce of this protein in peak I but not in peak II. Taken together, these
results suggest the presence in a soluble fraction from the kidney of
distinct methyl-accepting proteins, one of these being tentatively id
entified as the small G-protein CDC42.