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Two forms of the bovine brain Go that stimulate the inositol trisphosphate-mediated Cl- currents in Xenopus oocytes. Distinct guanine nucleotide binding properties
Ist Teil von
The Journal of biological chemistry, 1991-05, Vol.266 (15), p.9771-9777
Ort / Verlag
United States: American Society for Biochemistry and Molecular Biology
Erscheinungsjahr
1991
Quelle
MEDLINE
Beschreibungen/Notizen
Heterotrimeric GTP-binding proteins from bovine brain were resolved by fast protein liquid chromatography chromatography using
Mono Q columns. Two distinct forms of the protein Go were identified. Both forms had stochiometric amounts of alpha- and beta
gamma-subunits. The a-subunits of both forms were recognized by an alpha o-specific antiserum, but not by any of the alpha
i-specific antisera. The two forms showed distinct migration patterns on 9% sodium dodecyl sulfate-polyacrylamide gels containing
4-8 M urea gradients. Neither form comigrated with the recombinant alpha o1. Both the recombinant alpha o1 and the most abundant
form of Go were recognized by an antiserum, H-660, against a peptide encoding amino acids 3-17 of alpha i2. H-660 has been
shown previously to recognize alpha o and alpha i (Mumby, S. M., Pang, I. K., Gilman, A. G., and Sternweis, P. C. (1988) J.
Biol. Chem. 263, 2020-2026). This more abundant form is called Go A most likely corresponds to the cloned alpha o1. The less
abundant form, Go B, was not recognized by H-660. However, both forms of bovine brain Go were recognized by GC/2, an antiserum
against the N-terminal region of alpha o1. Hence alpha oA and alpha oB may be different in their N terminus regions. Neither
form of bovine brain Go was recognized by an antisera made to a peptide encoding the unique regions of the cloned alpha o2
from HIT cells (Hsu W. H., Rudolph, U., Sanford, J., Bertrand, P., Olate, J., Nelson, C., Moss, L.E., Boyd, A. E., III, Codina,
J., and Birnbaumer, L. (1990) J. Biol. Chem. 265, 11220-11226). Go A and Go B have similar guanine nucleotide binding and
release properties. Both release GDP within 1 min in the absence of added Mg2+. Both bind guanosine (GTP gamma S) rapidly
as well. However Go A binds GTP gamma S about 2.5-fold faster than Go B, in the absence of added Mg2+ ion. Both forms of Go
as well as the recombinant alpha o (alpha o1) can increase muscarinic stimulation of inositol trisphosphate-mediated Cl- current
in Xenopus oocytes. These data indicate that we have identified two structurally distinct forms of Go that have different
guanine nucleotide binding properties and are capable of functioning in the receptor-regulated phospholipase C pathway in
Xenopus oocytes.