tailieunhanh - Báo cáo khoa học: Changes in purine specificity in tandem GAF chimeras from cyanobacterial cyaB1 adenylate cyclase and rat phosphodiesterase 2

The C-terminal catalytic domains of the 11 mammalian phosphodiesterase families (PDEs) are important drug targets. Five of the 11 PDE families contain less well-characterized N-terminal GAF domains. cGMP is the lig-and for the GAF domains in PDEs 2, 5, 6 and 11, and cAMP is the ligand for PDE10. | ễFEBS Journal Changes in purine specificity in tandem GAF chimeras from cyanobacterial cyaBI adenylate cyclase and rat phosphodiesterase 2 Jurgen U. Linder Sandra Bruder Anita Schultz and Joachim E. Schultz Abteilung Pharmazeutische Biochemie Fakultat fur Chemie und Pharmazie Universitat Tubingen Germany Keywords adenylate cyclase cAMP cGMP cyclic nucleotide phosphodiesterase GAF domain Correspondence J. E. Schultz Abteilung Pharmazeutische Biochemie Fakultat fur Chemie und Pharmazie Universitat Tubingen Morgenstelle 8 72076 Tubingen Germany Fax 49 7071 295952 Tel 49 7071 2974676 E-mail Received 7 November 2006 revised 18 December 2006 accepted 12 January 2007 doi The C-terminal catalytic domains of the 11 mammalian phosphodiesterase families PDEs are important drug targets. Five of the 11 PDE families contain less well-characterized N-terminal GAF domains. cGMP is the ligand for the GAF domains in PDEs 2 5 6 and 11 and cAMP is the ligand for PDE10. Structurally related tandem GAF domains signalling via cAMP are present in the cyanobacterial adenylate cyclases cyaB1 and cyaB2. Because current high-resolution crystal structures of the tandem GAF domains of PDE2 and cyaB2 do not reveal how cNMP specificity is encoded we generated chimeras between the tandem GAF domains of cyaB1 and PDE2. Both bind the ligand in the GAF B subdomains. Segmental replacements in the highly divergent p1-P3 region of the GAF B subdomain of cyaB1 by the corresponding PDE2 regions switched signalling from cAMP to cGMP. Using 10 chimeric constructs we demonstrated that for this switch in purine specificity only 11 of the sequence of the cyanobacterial GAF B needs to be replaced by PDE2 sequences. We were unable however to switch the purine specificity of the PDE2 tandem GAF domain from cGMP to cAMP in reverse constructs . by replacement of PDE2 segments with those from the cyaB1 GAF tandem domain. The data provide a novel .