Soluble guanylate cyclase (sGC) is the primary mammalian nitric oxide (NO) sensor. Through the strong thermodynamic σ-trans effect of NO, binding of NO at the distal side of the ferrous heme induces cleavage of the proximal FeNHis bond, activating the catalytic domain of the enzyme. It has been proposed that nitroxyl (HNO) is also capable of activating sGC, but the key question remains as to whether HNO can induce cleavage of the FeNHis bond. Here we report calculated binding constants for 1-methylimidazole (MI) to [Fe(P)(X)] (P=porphine2−) where X=NO, HNO, CO, and MI to evaluate the trans interaction of these groups, X, with the proximal imidazole (histidine) in sGC. Systematic assessment of DFT methods suggests that the prediction of accurate MI binding constants is critically dependent on the inclusion of van der Waals interactions (−D functionals). Calculated (B3LYP-D/TZVP) MI binding constants for X=NO and MI are 110 and 5.6×105M−1, respectively, predicted only one order of magnitude higher than the corresponding experimentally determined values. MI binding constants where X=HNO and CO are consistently predicted to be essentially equal and ~six orders of magnitude larger than those of NO, indicating that CO and HNO mediate a weak thermodynamic trans effect in this system. Orbital analysis of the key σ-bonding orbital, π*h_dz2, and comparison of FeNMI bond lengths support this prediction. This suggests that HNO does not induce a σ-trans effect strong enough to promote cleavage of the FeNHis bond—a key step in the activation of sGC.