Extended Data Fig. 8: Structural analysis of Apyc1 and mechanism of cNMP degradation.
From: Phage anti-CBASS and anti-Pycsar nucleases subvert bacterial immunity
a, Structure guided multiple sequence alignment of Apyc1 proteins from the indicated phages or bacterial species and B. subtilis MBL phosphodiesterases. The strength of shading indicates degree of residue conservation. b, Overview of P. J14 Apyc1, P. xerothermodurans Apyc1, Bsp38 Apyc1, and B. subtilis YhfI crystal structures, with one monomer in colour and one monomer in grey. Detailed area highlights an Apyc1-specific loop that extends into the cNMP binding pocket. c, Detailed view of the residues coordinating the Zn2+ ions with cAMP modeled into the cNMP binding pocket. Numbers in parentheses indicate equivalent residue in SBSphiJ Apyc1. d, P. J14 Apyc1 crystallized in the presence of a hydrolysis-resistant phosphorothioate analog of cAMP resulted in clear phosphate and ribose density in the binding pocket and sparse density corresponding to the nucleobase. Polder omit map of cAMP contoured at 3.0 σ. e, TLC analysis of cCMP cleavage by SBSphiJ Apyc1 point mutants. f, HPLC analysis of Apyc1 cCMP and cUMP reaction products compared to synthesized 5′-CMP, 5′-UMP and 3′-CMP or 3′-UMP standards. Data are representative of 2 independent experiments.
