Fluoroaluminates activate transducin‐GDP by mimicking the γ‐phosphate of GTP in its binding site

Bigay, Joëlle; Déterre, Philippe; Pfister, Claude; Chabre, Marc

doi:10.1016/0014-5793(85)80004-1

Cited by 417 publications

(205 citation statements)

References 17 publications

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“…First, we prepared cytoplasmic chaperonin in its ADP-bound form and showed that it was fully competent for binary complex formation. Second, we prepared cytoplasmic chaperonin in an ADP-P, form by using beryllium fluoride complexes as a structural analog of Pi (4,5); this form of chaperonin, which mimicks the transitional (ADP-Pi) state that is equivalent to the ATPbound form, completely failed to bind unfolded target protein (Fig. 7).…”

Section: Discussionmentioning

confidence: 99%

Facilitated folding of actins and tubulins occurs via a nucleotide-dependent interaction between cytoplasmic chaperonin and distinctive folding intermediates.

Melki¹,

Cowan²

1994

Mol. Cell. Biol.

107

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In the cytoplasm of eukaryotes, the folding of actins and tubulins is facilitated via interaction with a heteromeric toroidal complex (cytoplasmic chaperonin). The folding reaction consists of the formation of a binary complex between the unfolded target protein and the chaperonin, followed by the ultimate release of the native polypeptide in an ATP-dependent reaction. Here we show that the mitochondrial chaperonin (cpn6O) and the cytoplasmic chaperonin both recognize a range of target proteins with different relative affinities; however, the cytoplasmic chaperonin shows the highest affinity for intermediates derived from unfolded tubulins and actins. These high-affinity actin and tubulin folding intermediates are distinct from the "molten globule" intermediates formed by noncytoskeletal target proteins in that they form relatively slowly. We show that the interaction between cytoplasmic chaperonin and unfolded target proteins depends on the chaperonin being in its ADP-bound state and that the release of the target protein occurs after a transition of the chaperonin to the ATP-bound state. Our data suggest a model in which ATP hydrolysis acts as a switch between conformational forms of the cytoplasmic chaperonin that interact either strongly or weakly with unfolded substrates.The biological function of most, if not all, proteins depends critically upon their three-dimensional structure. Although the information contained in the linear sequence of amino acids is thought to be sufficient. to specify this structure (2, 24), the correct folding of many proteins is not a spontaneous process under physiological conditions; rather, there is a requirement for interaction with a class of proteins or protein complexes known as molecular chaperones (reviewed in references 14, 18, and 39). In particular, one class of chaperones (typified by GroEL in prokaryotes) consist of multisubunit toroidal ring assemblies and are believed to function by providing a sequestered environment in which aberrant folding and aggregation are prevented; the correctly folded polypeptide is ultimately discharged in an Mg-ATP-dependent reaction. These toroidal structures are termed chaperonins (6,10,19,28,34,35,46). Actin and tubulin are the major soluble cytoplasmic proteins in eukaryotic cells and are the subunits from which actin filaments and microtubules are assembled. The folding of actins, tubulins, and actin-and tubulin-related proteins is facilitated by a chaperonin (15-17, 31, 49) that differs from its prokaryotic and mitochondrial homologs in that it is heteromeric: the multisubunit toroidal structure is assembled from eight different (though related) polypeptides (15, 38), one of which is the t-complex polypeptide TCP-1 (27, 42). As is the case for other chaperonins, the cytoplasmic chaperonin forms a stable binary complex with unfolded target polypeptides in the absence of Mg-ATP, and the hydrolysis of Mg-ATP is absolutely required for the generation of folded proteins (15-17). However, an important difference exists between the mecha...

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Section: Discussionmentioning

confidence: 99%

Facilitated folding of actins and tubulins occurs via a nucleotide-dependent interaction between cytoplasmic chaperonin and distinctive folding intermediates.

Melki¹,

Cowan²

1994

Mol. Cell. Biol.

107

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“…Al/NaF is able to activate all G proteins by substituting for the terminal phosphate in the presence of GDP to mimic GTP (27). NCDC, an inhibitor of phospholipase C (15,24), was then administered to the media in a cumulative manner (from 10 to 200 M).…”

Section: Methodsmentioning

confidence: 99%

Cellular mechanisms mediating rat renal microvascular constriction by angiotensin II.

Takenaka¹,

Suzuki²,

Fujiwara³

et al. 1997

J. Clin. Invest.

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To assess cellular mechanisms mediating afferent (AA) and efferent arteriolar (EA) constriction by angiotensin II (AngII), experiments were performed using isolated perfused hydronephrotic kidneys. In the first series of studies, AngII (0.3 nM) constricted AAs and EAs by 29 Ϯ 3 ( n ϭ 8, P Ͻ 0.01) and 27 Ϯ 3% ( n ϭ 8, P Ͻ 0.01), respectively. Subsequent addition of nifedipine restored AA but not EA diameter. Manganese (8 mM) reversed EA constriction by 65 Ϯ 9% ( P Ͻ 0.01). In the second group, the addition of N -ethylmaleimide (10 M), a Gi/Go protein antagonist, abolished AngIIinduced EA ( n ϭ 6) but not AA constriction ( n ϭ 6). In the third series of experiments, treatment with 2-nitro-4-carboxyphenyl-N , N -diphenyl-carbamate (200 M), a phospholipase C inhibitor, blocked both AA and EA constriction by AngII ( n ϭ 6 for each). In the fourth group, thapsigargin (1 M) prevented AngII-induced AA constriction ( n ϭ 8) and attenuated EA constriction (8 Ϯ 2% decrease in EA diameter at 0.3 nM AngII, n ϭ 8, P Ͻ 0.05). Subsequent addition of manganese (8 mM) reversed EA constriction. Our data provide evidence that in AAs, AngII stimulates phospholipase C with subsequent calcium mobilization that is required to activate voltage-dependent calcium channels. Our results suggest that AngII constricts EAs by activating phospholipase C via the Gi protein family, thereby eliciting both calcium mobilization and calcium entry. ( J.

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“…20,21 The increase in emission intensity which accompanies the activationdependent movement of the Switch II region into a hydrophobic pocket [1][2][3][4][5][6][7] provides a convenient and reliable method to assess the functional integrity of purified proteins. 22,23 Ga i contains two other Trp residues, W258 in the GTPase domain and W131 in the helical domain. Crystal structures show that the Trp in the helical domain of Ga i and Ga t proteins remains buried in both the inactive and active states, 3,4,6 and NMR studies have shown it to be refractory to activation.…”

mentioning

confidence: 99%

Trp fluorescence reveals an activation‐dependent cation‐π interaction in the Switch II region of Gα_i proteins

et al. 2009

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Crystal structures of Ga i (and closely related family member Ga t ) reveal much of what we currently know about G protein structure, including changes which occur in Switch regions. Ga t exhibits a low rate of basal (uncatalyzed) nucleotide exchange and an ordered Switch II region in the GDP-bound state, unlike Ga i , which exhibits higher basal exchange and a disordered Switch II region in Ga i GDP structures. Using purified Ga i and Ga t , we examined the intrinsic tryptophan fluorescence of these proteins, which reports conformational changes associated with activation and deactivation of Ga proteins. In addition to the expected enhancement in tryptophan fluorescence intensity, activation of GaGDP proteins was accompanied by a modest but notable red shift in tryptophan emission maxima. We identified a cation-p interaction between tryptophan and arginine residues in the Switch II of Ga i family proteins that mediates the observed red shift in emission maxima. Furthermore, amino-terminal myristoylation of Ga i resulted in a less polar environment for tryptophan residues in the GTPase domain, consistent with an interaction between the myristoylated amino terminus and the GTPase domain of Ga proteins. These results reveal unique insights into conformational changes which occur upon activation and deactivation of G proteins in solution.

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Fluoroaluminates activate transducin‐GDP by mimicking the γ‐phosphate of GTP in its binding site

Cited by 417 publications

References 17 publications

Facilitated folding of actins and tubulins occurs via a nucleotide-dependent interaction between cytoplasmic chaperonin and distinctive folding intermediates.

Facilitated folding of actins and tubulins occurs via a nucleotide-dependent interaction between cytoplasmic chaperonin and distinctive folding intermediates.

Cellular mechanisms mediating rat renal microvascular constriction by angiotensin II.

Trp fluorescence reveals an activation‐dependent cation‐π interaction in the Switch II region of Gα_i proteins

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Fluoroaluminates activate transducin‐GDP by mimicking the γ‐phosphate of GTP in its binding site

Cited by 417 publications

References 17 publications

Facilitated folding of actins and tubulins occurs via a nucleotide-dependent interaction between cytoplasmic chaperonin and distinctive folding intermediates.

Facilitated folding of actins and tubulins occurs via a nucleotide-dependent interaction between cytoplasmic chaperonin and distinctive folding intermediates.

Cellular mechanisms mediating rat renal microvascular constriction by angiotensin II.

Trp fluorescence reveals an activation‐dependent cation‐π interaction in the Switch II region of Gαi proteins

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About

Trp fluorescence reveals an activation‐dependent cation‐π interaction in the Switch II region of Gα_i proteins