The molecular mechanism of mammalian adenylyl cyclase activation has been elusive because there is as yet no structure representing a physiologically relevant basal, inactive state of the catalytic core (a C1:C2 heterodimer in the absence of activating ligands). The structure of the C2 homodimer of adenylyl cyclase has been used as a surrogate, but its conformation is probably influenced by the presence of two molecules of forskolin, which is used to stabilize the dimer interface, and the fact that it lacks a C1 domain, which is required for catalytic activity [6
The guanylyl cyclase structures provide new images of basal conformations that may be more physiologically relevant. The catalytic core of the Synechocystis
guanylyl cyclase enzyme [2
] assumes a conformation most similar to catalytically competent states of adenylyl cyclases [8
], although, surprisingly, it adopts a closed conformation that apparently needs to open before nucleotides can access the active site. This could represent a regulatory difference from the C. reinhardtii
guanylyl cyclase and mammalian cyclases. The C. reinhardtii
guanylyl cyclase [3
] adopts a conformation similar to that of the adenylyl cyclase C2 homodimer structure, in that the α1 helix, which binds to the β- and γ-phosphates of the nucleotide (Figure ), is not properly aligned with the rest of the active site. The structure may therefore serve as a useful model for the inactive state of mammalian cyclases. However, the fact that crystals could not be obtained without covalent modification of cysteines leaves open the possibility that this structure still does not represent the true ground-state conformation.
In 2005, Tews et al.
] described the active and inactive structures of a pH-sensing mycobacterial class III adenylyl cyclase. Importantly, this structure included the regulatory domains of the enzyme. In the active state, the catalytic domains assume the familiar wreath-like dimer. In the inactive state, they adopt a dramatically different conformation in which they interact extensively with the regulatory domains. Although this enzyme is more distantly related to mammalian adenylyl cyclases than the two guanylyl cyclases described here (approximately 23% identity) [2
], it highlights the fact that accessory domains, such as those found in the two guanylyl cyclase enzymes and in mammalian adenylyl cyclases, can have a dramatic impact on the conformation of the catalytic core in its inactive state.
There is also evidence that the ground-state structures of some class III nucleotide cyclases may consist simply of loosely associated, and therefore minimally active, catalytic domains. Although the C. reinhardtii
guanylyl cyclase enzyme [3
] was a homodimer under all conditions tested (J Winger, personal communication), the Cya2 catalytic core exists in an equilibrium between monomeric, dimeric and oligomeric forms [2
]. Forskolin and G proteins are known to dramatically enhance the affinity of the independently expressed C1 and C2 domains of mammalian adenylyl cyclase, and the Rv1900c adenylyl cyclase forms an asymmetric homodimer in its unliganded state, with an unusually open active-site cleft [12
Obviously, more structures of class III nucleotide cyclases are needed, especially of those that include regulatory domains. Although interpretation of the molecular mechanism of activation for nucleotide cyclases is somewhat hindered by the conditions required to bring about their crystallization, it is also possible that each class III enzyme has evolved a distinct basal conformation that can take optimal advantage of the regulatory inputs unique to that enzyme. This would probably enhance the fidelity of signaling when multiple cyclase isoforms are present.