scholarly journals Inhibition of NADPH oxidase activation by synthetic peptides mapping within the carboxyl-terminal domain of small GTP-binding proteins. Lack of amino acid sequence specificity and importance of polybasic motif.

1994 ◽  
Vol 269 (46) ◽  
pp. 29024-29031
Author(s):  
G Joseph ◽  
Y Gorzalczany ◽  
V Koshkin ◽  
E Pick
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Nadph Oxidase ◽  
Synthetic Peptides ◽  
Binding Proteins ◽  
Sequence Specificity ◽  
Gtp Binding Proteins ◽  
Gtp Binding ◽  
Carboxyl Terminal ◽  
Nadph Oxidase Activation

Molecular cloning of the cDNA for stimulatory GDP/GTP exchange protein for smg p21s (ras p21-like small GTP-binding proteins) and characterization of stimulatory GDP/GTP exchange protein

1991 ◽  
Vol 11 (5) ◽  
pp. 2873-2880
Author(s):  
K Kaibuchi ◽  
T Mizuno ◽  
H Fujioka ◽  
T Yamamoto ◽  
K Kishi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Exchange Reaction ◽  
Binding Proteins ◽  
Bovine Brain ◽  
Rat Tissues ◽  
Gtp Binding Proteins ◽  
Gtp Binding ◽  
Ras P21 ◽  
Exchange Protein

We have recently purified to near homogeneity the stimulatory GDP/GTP exchange protein for smg p21s (ras p21-like GTP-binding proteins) from bovine brain cytosol. This regulatory protein, named GDP dissociation stimulator (GDS), stimulates the GDP/GTP exchange reaction of smg p21s by stimulating the dissociation of GDP from and the subsequent binding of GTP to them. In this study, we have isolated and sequenced the cDNA of smg p21 GDS from a bovine brain cDNA library by using an oligonucleotide probe designed from the partial amino acid sequence of the purified smg p21 GDS. The cDNA has an open reading frame encoding a protein of 558 amino acids with a calculated Mr value of 61,066, similar to the Mr of 53,000 estimated for the purified smg p21 GDS by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and sucrose density gradient ultracentrifugation. The isolated cDNA is expressed in Escherichia coli, and the encoded protein exhibits smg p21 GDS activity. smg p21 GDS is overall hydrophilic, but there are several short hydrophobic regions. The smg p21 GDS mRNA is present in bovine brain and various rat tissues. smg p21 GDS has low amino acid sequence homology with the yeast CDC25 and SCD25 proteins, which may regulate the GDP/GTP exchange reaction of the yeast RAS2 protein, but not with ras p21 GTPase-activating protein, the inhibitory GDP/GTP exchange proteins (GDP dissociation inhibitor) for smg p25A and rho p21s, and the beta gamma subunits of heterotrimeric GTP-binding proteins such as Gs and Gi.

scholarly journals Molecular cloning of the cDNA for stimulatory GDP/GTP exchange protein for smg p21s (ras p21-like small GTP-binding proteins) and characterization of stimulatory GDP/GTP exchange protein.

1991 ◽  
Vol 11 (5) ◽  
pp. 2873-2880 ◽  
Author(s):  
K Kaibuchi ◽  
T Mizuno ◽  
H Fujioka ◽  
T Yamamoto ◽  
K Kishi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Exchange Reaction ◽  
Binding Proteins ◽  
Bovine Brain ◽  
Rat Tissues ◽  
Gtp Binding Proteins ◽  
Gtp Binding ◽  
Ras P21 ◽  
Exchange Protein

We have recently purified to near homogeneity the stimulatory GDP/GTP exchange protein for smg p21s (ras p21-like GTP-binding proteins) from bovine brain cytosol. This regulatory protein, named GDP dissociation stimulator (GDS), stimulates the GDP/GTP exchange reaction of smg p21s by stimulating the dissociation of GDP from and the subsequent binding of GTP to them. In this study, we have isolated and sequenced the cDNA of smg p21 GDS from a bovine brain cDNA library by using an oligonucleotide probe designed from the partial amino acid sequence of the purified smg p21 GDS. The cDNA has an open reading frame encoding a protein of 558 amino acids with a calculated Mr value of 61,066, similar to the Mr of 53,000 estimated for the purified smg p21 GDS by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and sucrose density gradient ultracentrifugation. The isolated cDNA is expressed in Escherichia coli, and the encoded protein exhibits smg p21 GDS activity. smg p21 GDS is overall hydrophilic, but there are several short hydrophobic regions. The smg p21 GDS mRNA is present in bovine brain and various rat tissues. smg p21 GDS has low amino acid sequence homology with the yeast CDC25 and SCD25 proteins, which may regulate the GDP/GTP exchange reaction of the yeast RAS2 protein, but not with ras p21 GTPase-activating protein, the inhibitory GDP/GTP exchange proteins (GDP dissociation inhibitor) for smg p25A and rho p21s, and the beta gamma subunits of heterotrimeric GTP-binding proteins such as Gs and Gi.

scholarly journals Antibodies directed against synthetic peptides distinguish between GTP-binding proteins in neutrophil and brain.

1987 ◽  
Vol 262 (30) ◽  
pp. 14683-14688 ◽  
Author(s):  
P Goldsmith ◽  
P Gierschik ◽  
G Milligan ◽  
C G Unson ◽  
R Vinitsky ◽  
...  
Keyword(s):  
Synthetic Peptides ◽  
Binding Proteins ◽  
Gtp Binding Proteins ◽  
Gtp Binding

scholarly journals Requirement for posttranslational processing of Rac GTP-binding proteins for activation of human neutrophil NADPH oxidase.

1993 ◽  
Vol 4 (3) ◽  
pp. 261-269 ◽  
Author(s):  
P G Heyworth ◽  
U G Knaus ◽  
X Xu ◽  
D J Uhlinger ◽  
L Conroy ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Human Neutrophil ◽  
Binding Proteins ◽  
Cysteine Residue ◽  
Posttranslational Processing ◽  
Nucleotide Exchange ◽  
Gtp Binding Proteins ◽  
Phagocyte Nadph Oxidase ◽  
Highly Active ◽  
Gtp Binding

Rac1 and Rac2 are closely related, low molecular weight GTP-binding proteins that have both been implicated in regulation of phagocyte NADPH oxidase. This enzyme system is composed of multiple membrane-bound and cytosolic subunits and when activated catalyzes the one-electron reduction of oxygen to superoxide. Superoxide and its highly reactive derivatives are essential for killing microorganisms. Rac proteins undergo posttranslational processing, primarily the addition of an isoprenyl group to a carboxyl-terminal cysteine residue. We directly compared recombinant Rac1 and Rac2 in a human neutrophil cell-free NADPH oxidase system in which cytosol was replaced by purified recombinant cytosolic components (p47-phox and p67-phox). Processed Rac1 and Rac2 were both highly active in this system and supported comparable rates of superoxide production. Under different cell-free conditions, however, in which suboptimal amounts of cytosol were present in the assay mixture, processed Rac2 worked much better than Rac1 at all but the lowest concentrations. This suggests that a factor in the cytosol may suppress the activity of Rac1 but not of Rac2. Unprocessed Rac proteins were only weakly able to support superoxide generation in either system, but preloading of Rac1 or Rac2 with guanosine 5'-O-(3-thio-triphosphate) (GTP gamma S) restored activity. These results indicate that processing is required for nucleotide exchange but not for interaction with oxidase components.

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