Structure And Catalytic Mechanism Of Arabidopsis Thaliana N~6-mAMP Deaminase ADAL

m~6A is the most abundant m RNA modification in eukaryotes with important biological function.N~6-methly-AMP is a proposed intermediate during m~6A-modified RNA metabolism,and will be incorporated into newly synthesized RNA by Pol II unless be properly turnover.AtADAL(Adenosine Deaminase-Like)is validated to undertake the function of converting N~6-m AMP into inosine monophosphate(IMP)to prevent it misuse.Although the function of ADAL has been reported,the three-dimensional structure of ADAL has not yet been determined.Here,we first solve the crystal structure of AtADAL in apo form,and reveal that the structure of ADAL shows TIM-barrel structure scaffolds,consisting of 9 parallel ?-strands and 15 peripheral-helices that surround the central ?-strands.We also found divalent metal ions Zn2+ are stably bound at the active site like other adenyl-deaminase family proteins do.To further elucidate the structure basis for substrate binding and catalytic activity of ADAL protein,we co-crystallize the Asn295(D295N),which is a key residue mutant at active site without catalytic activity,with GMP and IMP in the presence of Zn2+ and determined these complex structures.The structural comparisons between wild-type-AtADAL with D295N-IMP and D295N-GMP revealed that the ADAL in all these structures adopt almost identical conformation.Our further structural analysis have identified the substrate-binding pocket of AtADAL and compared it with that for adenosine deaminase(ADA),adenine deaminase(ADE)and AMP deaminase(AMPD)from multiple species.The comparisons reveal that plant ADAL1 may have the potential to catalyze different alkyl-group substituted substrate.Our studies of the crystal structure of AtADAL and its related complexes have shed lights on the molecular mechanism of the ADAL for substrates binding and catalyticactivity,and improved understanding of the metabolic pathway of m~6 A.Furthermore,given that the purine metabolism is involved in plenty of physiologic and pathologic contexts in human and highly conserved between arabidopsis thaliana and human,our studies provided the structural basis for drug design related to human ADAL.