Adenylate Cyclase in Heart and Skeletal Muscle

Drummond, George I.; Dunham, Jean; Nambi, Ponnal; Sano, M.

doi:10.1016/b978-0-08-023768-8.52358-6

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“…We found that phospholipase A treatment of microsomal fractions enhanced adenylate cyclase activity (Sulakhe & Dhalla, 1973), but recently we noted that such treatment caused inhibition of the sarcolemmal enzyme (Narayanan & Sulakhe, 1977). This observation suggested that microsomal and sarcolemmal enzyme respond differently to membrane-perturbing agents, despite the fact that the catalytic subunit of this enzyme present in these membranes displayed similarities in its kinetic behaviour (for example, affinities towards MgATP2and Mg2+) (Sulakhe & Dhalla, 1973; Louis & Drummond & Dunham, 1978). In the present study, we show that non-ionic detergents such as Lubrol-PX and Triton X-100 produce contrasting effects on the enzyme activity, depending on the membrane fraction used.…”

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confidence: 56%

“…The specific activities of cyclase of various membranes described here are comparable with those previously reported by us (Sulakhe & Dhalla, 1973;St. Louis & Sulakhe, 1976a,b;Narayanan & Sulakhe, 1977 and others (Drummond & Duncan, 1970;Drummond & Dunham, 1978;Entman et al, 1969;Katz et al, 1974).…”

Section: Resultsmentioning

confidence: 94%

“…In the present study, enzyme was assayed at 30°C to minimize inactivation caused by the non-ionic detergents, observed at the higher assay temperature (37°C) (Drummond & Dunham, 1978;N. Narayanan & P. V. Sulakhe, unpublished work).…”

Section: Stimulation Of the Solubilized Enzyme By P[nh]ppgmentioning

confidence: 99%

“…In view of these and other observations described here, it appears that adenylate cyclase is a component of sarcoplasmic reticulum (microsomal particles), although the content of this enzyme is much greater in heart sarcolemma. Drummond & Dunham (1978) reported that Lubrol-PX and Triton X-100 stimulated adenylate cyclase activity of 1000OOg particles (microsomal fraction) and inhibited the enzyme present in washed particles isolated from rabbit heart ventricles. Intheir…”

Section: Vol 175mentioning

confidence: 99%

“…Localization of adenylate cyclase (EC 4.6.1.1) in heart has been a controversial subject. Although earlier studies (Entman et al, 1969;Dhalla et al, 1970;Sulakhe & Dhalla, 1973;Katz et al, 1974) reported the presence of this enzyme activity in microsomal fraction of cardiac muscle, the recovery of the enzyme was low (at best 3-5% of the homogenate activity) (Sulakhe & Dhalla, 1973;Drummond & Dunham, 1978) and therefore it has been suggested that contamination of microsomal fraction by sarcolemma would account for this enzyme activity in microsomal fractions (Besch et al, 1976;Engelhard et al, 1976). This view is strengthened by the observations that the sarcoplasmic-reticulum-enriched microsomal fraction also contains sarcolemmal membranes (Besch etal., 1976;Jones etal., 1977) and that purified sarcolemma possesses highly active adenylate cyclase activity recovered in good yield (20-40%) (Hui et al, 1976;Sulakhe et al, 1976a; Louis & .…”

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confidence: 94%

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Differential effects of non-ionic detergents on microsomal and sarcolemmal adenylate cyclase in cardiac muscle

Sulakhe

Narayanan

1978

Biochemical Journal

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1. About 4 and 23% of the homogenate adenylate cyclase activity was recovered in the microsomal and sarcolemmal fractions isolated from guinea-pig heart ventricles. 2. Cardiac microsomal adenylate cyclase activity [basal as well as p[NH]ppG (guanyl-5'-yl imidodiphosphate)- and NaF-stimulated] was increased over 2-fold in the presence of Lubrol-PX (0.01-0.1%). 3. The sarcolemmal enzyme, however, showed concentration-dependent inhibition caused by the detergent under all assay conditions, except when p[NH]ppG was included in the assay. In the latter case, the detergent (0.01-0.02%) caused a modest increase (30-45%) in enzyme activity. 4. Another non-ionic detergent, Triton X-100, also stimulated the microsomal cyclase and inhibited the sarcolemmal enzyme. 5. With either membrane fraction, Lubrol-PX solubilized the enzyme when the detergent/membrane protein ratio was 2.5 (mumol of detergent/mg of protein). 6. The findings with homogenate and a washed particulate fraction resembled those obtained with sarcolemma, and those with isolated sarcoplasmic reticulum resembled those with microsomal preparations. 7. p[NH]ppG, and to some extent NaF, protected the detergent-induced inactivation of the enzyme observed at higher detergent concentrations (0.5% Lubrol-PX and 0.05-0.5% Triton X-100). 8. In the absence of detergents, p[NH]ppG increased the basal enzyme activity about 2-fold in microsomal fractions, but did not appreciably stimulate the sarcolemmal enzyme. Isoproterenol, on the other hand, increased the sarcolemmal enzyme activity (>2-fold) in the presence of p[NH]ppG and caused only moderate stimulation (31%) of the microsomal enzyme under these conditions. 9. These findings support the view that, although the bulk of adenylate cyclase resides in heart sarcolemma (plasma membrane), the microsomal activity cannot be accounted for solely by contamination of the microsomal fraction with sarcolemma, as has been suggested by others [Besch, Jones & Watanabe (1976) Circ. Res.39, 586-595; Engelhard, Plut & Storm (1976) Biochim. Biophys. Acta451, 48-61]. Further, the results of this study show that cardiac sarcoplasmic-reticulum membranes possess this enzyme.

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confidence: 56%