Abstract
Mutational replacements of specific residues in the GTP-binding pocket of the 21-kDa ras proteins (p21(ras)) reduce their GTPase activity. To test the possibility that the cognate regions of G protein α chains participate in GTP binding and hydrolysis, we compared signaling functions of normal and mutated α chains (termed α(s)) of G(s), the stimulatory regulator of adenylyl cyclase. α(s) chains were expressed in an α(s)-deficient S49 mouse lymphoma cell line, cyc- α(s) in which leucine replaces glutamine 227 (corresponding to glutamine 61 of p21(ras)) constitutively activates adenylyl cyclase and reduces the k(cat) for GTP hydrolysis more than 100-fold. There is a smaller reduction in GTPase activity in another mutant in which valine replaces glycine 49 (corresponding to glycine 12 of p21(ras)). This mutant α(s) is a poor activator of adenylyl cyclase. Moreover, the glycine 49 protein, unlike normal α(s), is not protected against tryptic cleavage by hydrolysis resistant GTP analogs; this finding suggests impairment of the mutant protein's ability to attain the active (GTP-bound) conformation. We conclude that α(s) residues near glutamine 227 and glycine 49 participate in binding and hydrolysis of GTP, although the GTP binding regions of α(s) and p21(ras) are not identical.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 15467-15474 |
| Number of pages | 8 |
| Journal | Journal of Biological Chemistry |
| Volume | 264 |
| Issue number | 26 |
| State | Published - 1989 |
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ASJC Scopus subject areas
- Biochemistry
Cite this
Mutations in the GTP-binding site of G(sα) alter stimulation of adenylyl cyclase. / Masters, S. B.; Miller, R. T.; Chi, M. H.; Chang, F. H.; Beiderman, B.; Lopez, N. G.; Bourne, H. R.
In: Journal of Biological Chemistry, Vol. 264, No. 26, 1989, p. 15467-15474.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Mutations in the GTP-binding site of G(sα) alter stimulation of adenylyl cyclase
AU - Masters, S. B.
AU - Miller, R. T.
AU - Chi, M. H.
AU - Chang, F. H.
AU - Beiderman, B.
AU - Lopez, N. G.
AU - Bourne, H. R.
PY - 1989
Y1 - 1989
N2 - Mutational replacements of specific residues in the GTP-binding pocket of the 21-kDa ras proteins (p21(ras)) reduce their GTPase activity. To test the possibility that the cognate regions of G protein α chains participate in GTP binding and hydrolysis, we compared signaling functions of normal and mutated α chains (termed α(s)) of G(s), the stimulatory regulator of adenylyl cyclase. α(s) chains were expressed in an α(s)-deficient S49 mouse lymphoma cell line, cyc- α(s) in which leucine replaces glutamine 227 (corresponding to glutamine 61 of p21(ras)) constitutively activates adenylyl cyclase and reduces the k(cat) for GTP hydrolysis more than 100-fold. There is a smaller reduction in GTPase activity in another mutant in which valine replaces glycine 49 (corresponding to glycine 12 of p21(ras)). This mutant α(s) is a poor activator of adenylyl cyclase. Moreover, the glycine 49 protein, unlike normal α(s), is not protected against tryptic cleavage by hydrolysis resistant GTP analogs; this finding suggests impairment of the mutant protein's ability to attain the active (GTP-bound) conformation. We conclude that α(s) residues near glutamine 227 and glycine 49 participate in binding and hydrolysis of GTP, although the GTP binding regions of α(s) and p21(ras) are not identical.
AB - Mutational replacements of specific residues in the GTP-binding pocket of the 21-kDa ras proteins (p21(ras)) reduce their GTPase activity. To test the possibility that the cognate regions of G protein α chains participate in GTP binding and hydrolysis, we compared signaling functions of normal and mutated α chains (termed α(s)) of G(s), the stimulatory regulator of adenylyl cyclase. α(s) chains were expressed in an α(s)-deficient S49 mouse lymphoma cell line, cyc- α(s) in which leucine replaces glutamine 227 (corresponding to glutamine 61 of p21(ras)) constitutively activates adenylyl cyclase and reduces the k(cat) for GTP hydrolysis more than 100-fold. There is a smaller reduction in GTPase activity in another mutant in which valine replaces glycine 49 (corresponding to glycine 12 of p21(ras)). This mutant α(s) is a poor activator of adenylyl cyclase. Moreover, the glycine 49 protein, unlike normal α(s), is not protected against tryptic cleavage by hydrolysis resistant GTP analogs; this finding suggests impairment of the mutant protein's ability to attain the active (GTP-bound) conformation. We conclude that α(s) residues near glutamine 227 and glycine 49 participate in binding and hydrolysis of GTP, although the GTP binding regions of α(s) and p21(ras) are not identical.
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UR - http://www.scopus.com/inward/citedby.url?scp=0024468792&partnerID=8YFLogxK
M3 - Article
C2 - 2549064
AN - SCOPUS:0024468792
VL - 264
SP - 15467
EP - 15474
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 26
ER -