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  • br Acknowledgments This review is based

    2021-09-15


    Acknowledgments This review is based in part on research conducted with financial support from the General Research Fund (17112914) of the Hong Kong Research Grant Council and from the Seed Funding Programme for Basic Research by University Research Committee of the University of Hong Kong (201511159163). The authors would like to thank Ms. Yee Har Chung and Mr. Godfrey Man for technical assistance and Ms. Ivy Wong for editorial assistance.
    Introduction Cyclic GMP was first purified and identified in rat urine in 1963 by Ashman and colleagues [1]. The enzymes that catalyze the conversion of GTP into cGMP were discovered 6years later by three separate groups [2], [3], [4]. The cGMP synthesizing activity was initially called guanyl cyclase, then guanylate cyclase and in recent literature it is most often referred to as guanylyl cyclase. The Nomenclature Committee of the International Union of Biochemistry and Molecular Biology indicates that the accepted name is guanylate cyclase or GTP pyrophosphate-lyase, EC 4.6.1.2. The reaction catalyzed is: divalent metal bound GTP→cGMP+PPi. The divalent metal cofactor under biologic conditions is Mg2+, but many early investigations used Mn2+ in the presence of non-ionic detergent in order to artificially stimulate the enzymes because the biologic activators of the respective enzymes were not known. As a result of these initial studies, it was determined that the enzyme could be separated into distinct soluble and particular forms with unique kinetic properties and tissue distributions [5], [6]. However, the true Wiskostatin of the family was not fully appreciated until molecular cloning identified cDNAs that encoded individual enzymes. Mammals express four soluble (α1, α2, β1 and β2) guanylyl cyclase subunits and seven bona fide single membrane-spanning forms known as GC-A, GC-B, GC-C, GC-D, GC-E, GC-F and GC-G (see Table 1 for ligands and “knockout” phenotypes associated with each Wiskostatin cyclase). However, GC-D and GC-G are pseudogenes in humans [7]. The soluble forms exist as heterodimers and the transmembrane members are homodimers, except for GC-C, which appears to be a homotrimer. The minimal guanylyl cyclase catalytic unit is a dimer [8], [9]. The soluble forms contain an amino terminal heme-binding, dimerization and carboxyl-terminal catalytic domains. The membrane-spanning forms contain an extracellular ligand-binding, transmembrane, kinase homology, dimerization and carboxyl-terminal catalytic domains (see Fig. 1 for topology of each cyclase). The latter domain is the most conserved between family members and is homologous to the catalytic domain of adenylyl cyclase [10], [11]. Interestingly, Caenorhabdiis elegans are predicted to have at least 26 membrane-spanning and 7 soluble guanylyl cyclases based on genome analysis [12], but humans only express five membrane spanning and 3 soluble subunits, which may indicate the human forms are more pleotropic than the enzymes found in worms. Multimembrane-spanning guanylyl cyclases similar to adenylyl cyclases were identified in Dictyostelium discoidium, Plasmodium falciparum, Paramecium tetraurelia and Tetrahymena pyriformis. These guanylyl cyclases probably evolved from adenylyl cyclase based upon topology [13]. One guanylyl cyclase was identified in cyanobacteria [14] and the crystal structure of the catalytic domain of this enzyme was solved [15]. Additionally, the crystal structure of an inactive soluble guanylyl cyclase from the green algae Chlamydomonas reinhardtii was solved [16]. According to the structures described above, the guanylyl cyclase catalytic domain contains a central seven-stranded β sheet surrounded by several α helices in a wreath-like structure that is characteristic of class III nucleotide cyclases. The central cleft of the momodimeric form is at the dimer interface and contains two symmetric active sites, which is consistent with positive cooperative substrate-velocity curves observed for particulate cyclases [15]. Each monomer contributes unique residues to the catalytic site. Joubert and colleagues created a mutant form of the catalytic domain of rat GC-A by making single amino acid substitutions on each monomer to inactive only one GTP binding site and observed linear kinetics, consistent with mammalian GCs adopting a similar structure to the cyanobacterial GC [17]. Mutagenesis studies converting human GC-E and rat soluble guanylyl cyclase to adenylyl cyclases are consistent with mammalian guanylyl cyclases adopting similar structures and catalytic mechanisms as adenylyl cyclases [10], [11]. No structure of an active mammalian guanylyl cyclase has been solved to date.