An EFI example from the Computation Core and AH Bridging Project showing the ability to go from sequence to function for a target from Pseudoalteromonas atlantica T6c which did not have an experimental crystal structure. Homology models and docking predicted adenine and various adenosine derivatives within the active site. The AH Bridging Project verified that the enzyme catalyzes hydrolysis of N-6-substituted adenine derivatives, several of which are cytokinins
An enzyme of unknown function within the amidohydrolase superfamily was discovered to catalyze the hydrolysis of N-6-substituted adenine derivatives, several of which are cytokinins. Cytokinins are a common type of plant hormone and N-6-substituted adenines are also found as modifications to tRNA. Patl2390, from Pseudoalteromonas atlantica T6c, was shown to hydrolytically deaminate N-6-isopentenyladenine to hypoxanthine and isopentenylamine with a k(cat)/K(m) of 1.2 × 10(7) M(-1) s(-1). Additional substrates include N-6-benzyl adenine, cis- and trans-zeatin, kinetin, O-6-methylguanine, N-6-butyladenine, N-6-methyladenine, N,N-dimethyladenine, 6-methoxypurine, 6-chloropurine, and 6-thiomethylpurine. This enzyme does not catalyze the deamination of adenine or adenosine. A comparative model of Patl2390 was computed using the three-dimensional crystal structure of Pa0148 (PDB code 3PAO ) as a structural template, and docking was used to refine the model to accommodate experimentally identified substrates. This is the first identification of an enzyme that will hydrolyze an N-6-substituted side chain larger than methylamine from adenine.
Figure 1. Sequence similarity network created using Cytoscape(6) of cog1816 from the amidohydrolase superfamily. Each node in the network represents a single sequence, and each edge (depicted as lines) represents the pairwise connection between two sequences at a BLAST E-value of better than 1 × 10–70. Lengths of edges are not significant, except for tightly clustered groups, which are more closely related than sequences with only a few connections. Group 5 contains the E. coli adenosine deaminase. The yellow dots in Group 3 represent proteins that will deaminate adenine. The proteins represented by the pink dots at the periphery of Group 3 are characterized in this paper.
2011 AH Superfamily Publication
Reprinted with permission from ACS Chemical Biology. © 2011 American Chemical Society.