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PMID:23968233
| Citation |
Hitchcock, DS, Fan, H, Kim, J, Vetting, M, Hillerich, B, Seidel, RD, Almo, SC, Shoichet, BK, Sali, A and Raushel, FM (2013) Structure-guided discovery of new deaminase enzymes. J. Am. Chem. Soc. 135:13927-33 |
|---|---|
| Abstract |
A substantial challenge for genomic enzymology is the reliable annotation for proteins of unknown function. Described here is an interrogation of uncharacterized enzymes from the amidohydrolase superfamily using a structure-guided approach that integrates bioinformatics, computational biology, and molecular enzymology. Previously, Tm0936 from Thermotoga maritima was shown to catalyze the deamination of S-adenosylhomocysteine (SAH) to S-inosylhomocysteine (SIH). Homologues of Tm0936 homologues were identified, and substrate profiles were proposed by docking metabolites to modeled enzyme structures. These enzymes were predicted to deaminate analogues of adenosine including SAH, 5'-methylthioadenosine (MTA), adenosine (Ado), and 5'-deoxyadenosine (5'-dAdo). Fifteen of these proteins were purified to homogeneity, and the three-dimensional structures of three proteins were determined by X-ray diffraction methods. Enzyme assays supported the structure-based predictions and identified subgroups of enzymes with the capacity to deaminate various combinations of the adenosine analogues, including the first enzyme (Dvu1825) capable of deaminating 5'-dAdo. One subgroup of proteins, exemplified by Moth1224 from Moorella thermoacetica, deaminates guanine to xanthine, and another subgroup, exemplified by Avi5431 from Agrobacterium vitis S4, deaminates two oxidatively damaged forms of adenine: 2-oxoadenine and 8-oxoadenine. The sequence and structural basis of the observed substrate specificities were proposed, and the substrate profiles for 834 protein sequences were provisionally annotated. The results highlight the power of a multidisciplinary approach for annotating enzymes of unknown function. |
| Links |
PubMed PMC3827683 Online version:10.1021/ja4066078 |
| Keywords |
Catalytic Domain; Crystallography, X-Ray; Enzyme Assays; Kinetics; Models, Molecular; Molecular Structure; Nucleoside Deaminases/chemistry; Nucleoside Deaminases/metabolism; Structure-Activity Relationship; Substrate Specificity |
| edit table |
Significance
Annotations
| Gene product | Qualifier | GO Term | Evidence Code | with/from | Aspect | Extension | Notes | Status |
|---|---|---|---|---|---|---|---|---|
|
enables |
GO:0050270: S-adenosylhomocysteine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Seeded From UniProt |
complete | |||
| GO:0050270: S-adenosylhomocysteine deaminase activity |
ECO:0000314: |
F |
Table 1, row m, kcat/Km value for SAH |
complete | ||||
| GO:0050270: S-adenosylhomocysteine deaminase activity |
ECO:0000314: |
F |
Table 1, row a, kcat/Km value for SAH |
complete | ||||
|
enables |
GO:0050270: S-adenosylhomocysteine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Seeded From UniProt |
complete | |||
| GO:0004000: adenosine deaminase activity |
ECO:0000314: |
F |
Table 1, row c, kcat/Km value for Ado |
complete | ||||
|
enables |
GO:0004000: adenosine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Seeded From UniProt |
complete | |||
| GO:0004000: adenosine deaminase activity |
ECO:0000314: |
F |
Table 1, row i, kcat/Km value for Ado |
complete | ||||
|
enables |
GO:0004000: adenosine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Seeded From UniProt |
complete | |||
| GO:0090614: 5'-methylthioadenosine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Table 1 shows kinetic paramters evaluated for different substrates. The annotation is made for the substrate for which highest activity is detected. This protein is Xcc2270. |
complete | ||||
| GO:0090614: 5'-methylthioadenosine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Table 1 shows the kinetic parameters for various substrates. The annotation is for the substrate for which the enzyme shows highest activity. This enzyme is Pa3170 |
complete | ||||
| GO:0090613: 5'-deoxyadenosine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Table 1 shows kinetic parameters evaluated for various substrates. this enzyme is Dvu1825. The annotation is for that substrate for which the enzyme shows greatest activity. |
complete | ||||
|
enables |
GO:0050270: S-adenosylhomocysteine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Seeded From UniProt |
complete | |||
| GO:0050270: S-adenosylhomocysteine deaminase activity |
ECO:0000314: |
F |
Table 1 shows the kinetic parameters for variiuos substrates. The annotation is made for the substrate for which the enzyme shows the highest activity. |
complete | ||||
|
enables |
GO:0050270: S-adenosylhomocysteine deaminase activity |
ECO:0000314: direct assay evidence used in manual assertion |
F |
Seeded From UniProt |
complete | |||
Notes
See also
References
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