Fisher R, Blumenthal T
J Biol Chem. 1982 Feb 25;257(4):1702-4.
Alterations in RNA polymerase structure can be detected using initial trypsin cleavage rates as a conformational probe. Both template (poly[d(A-T) . d(A-T)] and the RNA polymerase inhibitor, heparin, alter the rates at which the subunits of the enzyme are cleaved. However, while the presence of poly[d(A-T) . d(A-T)] slows the cleavage of subunits beta, sigma, and alpha by trypsin, heparin accelerates the cleavage of beta and sigma. Furthermore, the presence of heparin does not prevent the effect of poly[d(A-T) . d(A-T)] on the beta and sigma cleavage rates. Thus, heparin does not eliminate the interaction between DNA and RNA polymerase. That heparin does alter the nature of this interaction is demonstrated by the fact that template decreases the trypsin cleavage rate of subunit alpha in the absence, but not in the presence, of heparin. Like heparin, the addition of RNA to the reaction increases the accessibility of beta and sigma to trypsin. Hence the interaction of heparin with RNA polymerase may mimic the product, rather than the template, interaction.
利用初始胰蛋白酶切割速率作为构象探针，可以检测RNA聚合酶结构的改变。模板（聚[d(A-T).d(A-T)]）和RNA聚合酶抑制剂肝素都会改变酶亚基的切割速率。然而，虽然聚[d(A-T).d(A-T)]的存在会减缓胰蛋白酶对β、σ和α亚基的切割，但肝素会加速β和σ的切割。此外，肝素的存在并不妨碍聚[d(A-T).d(A-T)]对β和σ切割速率的影响。因此，肝素不会消除DNA与RNA聚合酶之间的相互作用。肝素确实改变了这种相互作用的性质，这一事实表明，在没有肝素的情况下模板会降低α亚基的胰蛋白酶切割速率，但在有肝素的情况下则不会。与肝素一样，向反应中添加RNA会增加β和σ对胰蛋白酶的可及性。因此，肝素与RNA聚合酶的相互作用可能模拟了产物而非模板的相互作用。
