Dimerization region of soluble guanylate cyclase characterized by bimolecular fluorescence complementation in vivo

The ubiquitously expressed nitric oxide (NO) receptor soluble guanylate cyclase (sGC) plays a key role in signal transduction. Binding of NO to the N-terminal prosthetic heme moiety of sGC results in an approximate 200-fold activation of the enzyme and an increased conversion of GTP into its second messenger cyclic GMP (cGMP). sGC exists as an heterodimer whose dimerization is mediated mainly by the central region of the enzyme. In the present work, we constructed deletion mutants within the predicted dimerization region of the sGC alpha1- and beta1-subunit to precisely map the sequence segments crucial for subunit dimerization. To track mutation-induced alterations of sGC dimerization we used a bimolecular fluorescence complementation (BiFC) approach which allows visualizing sGC heterodimerization in a non-invasive manner in living cells. Our study suggests that segments spanning amino acids alpha1-363-372, alpha1-403-422, alpha1-440-459 and beta1-212-222, beta1-304-333, beta1-344-363, beta1-381-400 within the predicted dimerization region are involved in the process of heterodimerization and therefore in the expression of functional sGC.