Guanylate Cyclase: Regulation of Cyclic GMP Metabolism

Mittal, Chandra K.; Murad, Ferid

doi:10.1007/978-3-642-68111-0_5

Cited by 46 publications

(26 citation statements)

References 154 publications

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“…In addition, the activity of the enzyme was monitored in the presence of Mn2+ and Mg2+. since it had been suggested that activation of guanylate cyclase during purification is accompanied by an increase in the Mg2'/Mn2+ activity ratio and by a decrease in stimulation by NO or sodium nitroprusside [2,9]. The Mg2+/Mn2+ activity ratio determined in the absence of sodium nitroprusside varied considerably during the purification.…”

Section: Comments On Purificationmentioning

confidence: 99%

Purifieation of a Soluble, Sodium‐Nitroprusside‐Stimulated Guanylate Cyclase from Bovine Lung

Gerzer

Hofmann

Schultz

1981

European Journal of Biochemistry

140

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A soluble, sodium‐nitroprusside‐stimulated guanylate cyclase has been purified from bovine lung by DEAE cellulose chromatography, ammonium sulfate precipitation, chromatography on Blue Sepharose CL‐6B and preparative gel electrophoresis. Apparent homogeneity was obtained after at least 7000‐fold purification with a yield of 3%. A single stained band (Mr 72000) was observed after gel electrophoresis in the presence of sodium dodecyl sulfate. The purified enzyme migrated as one band also under non‐denaturing conditions in acrylamide gels (5–12%). The mobility of this band corresponded to an Mr of 145000. The enzyme sedimented on sucrose gradients with an S20.w of 7.0 S. Gel filtration yielded a Stokes' radius of 4.6 nm. These data suggest that the enzyme has an Mr of approximately 150000 and consists of two, presumably identical, subunits of Mr 72000. Sodium nitroprusside stimulated the purified enzyme 15‐fold and 140‐fold to specific activities of 8.5 and 15.7 pmol cGMP formed min−1 mg−1 in the presence of Mn2+ and Mg2+, respectively. Formation of cGMP was proportional to the incubation time and to the amount of enzyme added. The stimulatory effect of sodium nitroprusside was half‐maximal at about 2 μM. was observed immediately after addition and could be reversed either by dilution or by removal of sodium nitroprusside on a Sephadex G‐25 column. The purified enzyme in the absence of catalase was stimulated by sodium nitroprusside, N‐methyl‐N′‐nitro‐N‐nitrosoguanidine and 3‐morpholino‐sydnonimine and in the presence of catalase by sodium nitrite and sodium azide. In the presence of Mn2+ and sodium nitroprusside, the purified enzyme catalyzed the formation of cAMP from ATP at a rate of 0.6 μmol min−1 mg−1.

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Section: Comments On Purificationmentioning

confidence: 99%

Purifieation of a Soluble, Sodium‐Nitroprusside‐Stimulated Guanylate Cyclase from Bovine Lung

Gerzer

Hofmann

Schultz

1981

European Journal of Biochemistry

140

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“…In most tissues, at least two forms of guanylyl cyclases (GTP pyrophosphate-lyase (cyclizing); EC 4.6.1.2) exist; the soluble and the plasma membranebound enzyme forms differ in structure, regulation and other properties [1,2]. Their enzymatic product, cGMP, acts as an intracellular messenger, activating cGMPdependent protein kinases [3] and regulating cGMPsensitive ion channels [4].…”

Section: Introductionmentioning

confidence: 99%

“…The plasma membrane-bound forms of guanylyl cyclase known so far exist as single polypeptide chains and are regulated by peptide hormones binding to an extracellular receptor domain of the protein [5]. On the other hand, the soluble enzyme form is a heterodimer, which is regulated by nitric oxide (NO) and NO-containing compounds [1,2]. This enzyme, which consists of two subunits, has been purified by several laboratories to apparent homogenity; the reported Mr of the larger subunit varied between 73 and 82 kDa on SDSpolyacrylamide gels [13,14].…”

Section: Introductionmentioning

confidence: 99%

The primary structure of the larger subunit of soluble guanylyl cyclase from bovine lung Homology between the two subunits of the enzyme

et al. 1990

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The primary structure of the larger subunit of the soluble guanylyl cyclase from bovine lung, which catalyzes the formation of cyclic GMP from GTP, has been determined. Two clones, isolated from two bovine libraries yielded a total of 3261 bp with a coding region of 2073 bp. The open reading frame encodes a protein of 691 amino acids and a molecular mass of 77 500. The deduced amino acid sequence reveals regions which are, to a large extent, homologous to the sequence of the smaller subunit of the enzyme as well as to the sequences of other gyanylyl and adenylyl cyclases.

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“…2). If Mg was omitted, none of the patches exhibited a current, suggesting that the Mg-GTP-induced current derived from the synthesis of cGMP by guanylate cyclase, which requires Mg for enzymatic activity (13). The light-evoked changes in the current induced by cGMP or by MgGTP were similar ( Figs.…”

Section: Resultsmentioning

confidence: 90%

Excised patches of plasma membrane from vertebrate rod outer segments retain a functional phototransduction enzymatic cascade.

Ertel

1990

Proc. Natl. Acad. Sci. U.S.A.

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Ion channels in excised patches of plasma membrane are generally considered to be isolated from any intracellular regulation mechanisms. For example, in excised patches of vertebrate rod outer segment plasma membrane, the cGMP-activated cation channels have traditionally been studied in room light because the enzyme cascade linking photon absorption to channel closure was assumed to be inoperative. To investigate the possibility that, in fact, such excised patches retain a functional phototransduction enzymatic cascade, this same preparation was studied in darkness. Patches excised in the dark were found to retain the light sensitivity of their cGMP-induced conductance and the ability to synthesize cGMP. In the presence of guanosine 5'-[y-thioltriphosphate (GTP[yS]), a nonhydrolyzable GTP analog, light suppresses the cGMP-induced conductance irreversibly. Furthermore, inhibitors of phosphodiesterase activity reduce light sensitivity, whereas activated phosphodiesterase or activated transducin does not directly affect the channels. These results (i) establish that excised patches from rod outer segment retain functional phototransduction enzymes, (it) support the classical view that channel opening is modulated by phosphodiesterase-mediated cGMP hydrolysis, and, most surprisingly, (i) demonstrate that diffusion in excised patches is so restricted that local enzymes can induce variations in the concentration of small molecules. The indication that excised patches are not as simple as usually surmised opens the possibility of using them to study other intracellular transduction mechanisms.In the traditional representation of excised patches, membrane proteins are considered to be isolated from any intracellular regulation mechanism involving either soluble enzymes or diffusible messengers. This isolation is presumed to arise from two phenomena: (i) soluble enzymes wash out from the patch when it is excised from the rest of the cell, and (ii) any intracellular messengers that might be synthesized by membrane enzymes diffuse instantaneously out into the "infinite" volume of the perfusion bath. In most cases, however, no attempt has been made to investigate the validity of these assumptions. The work reported here examines this problem in the specific case of excised patches of vertebrate rod outer segment (ROS) plasma membrane.Vertebrate vision is initiated in photoreceptor outer segments by a process that links the absorption of photons to the closure of ion channels in the plasma membrane (1,2 The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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Guanylate Cyclase: Regulation of Cyclic GMP Metabolism

Cited by 46 publications

References 154 publications

Purifieation of a Soluble, Sodium‐Nitroprusside‐Stimulated Guanylate Cyclase from Bovine Lung

Purifieation of a Soluble, Sodium‐Nitroprusside‐Stimulated Guanylate Cyclase from Bovine Lung

The primary structure of the larger subunit of soluble guanylyl cyclase from bovine lung Homology between the two subunits of the enzyme

Excised patches of plasma membrane from vertebrate rod outer segments retain a functional phototransduction enzymatic cascade.

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