Protein Variants | Comment | Organism |
---|---|---|
D147A | site-directed mutagenesis, the mutant shows a decrease in activity compared to the wild-type enzyme | Bos taurus |
E138A | site-directed mutagenesis, the mutation causes a modest increase in the rate of NAD+ transformation which is proportional to its concentration. At 4.0 M, the rate increase is about 1.2fold and the formation of beta-1'-O-methyl ADP-ribose amounts to about 80% of the total reaction products. The observed selectivity in favor of methanolysis is similar to that of wild-type enzyme. The ADP-ribosyl cyclase activity of E138A mutant is more affected by the competing nucleophile, i.e. formation of ADP-ribose and cADPR are reduced by 75% and 90% respectively at 4.0 M methanol, the mutant shows an increase in ADP cyclization and higly reduced overall activity compared to the wild-type enzyme | Bos taurus |
E138Q | site-directed mutagenesis, in the presence of methanol, mutant E138Q efficiently catalyzes the formation of beta-1'-O-methyl ADP-ribose. But in contrast with mutant E138A, and like the wild-type enzyme, solvolysis does not affect the overall turnover rate of NAD+ indicating that the formation of the E.ADP-ribosyl intermediate is still rate limiting | Bos taurus |
E218A | site-directed mutagenesis, the mutant shows a decrease in activity compared to the wild-type enzyme | Bos taurus |
E218Q | site-directed mutagenesis, the mutant shows a decrease in activity compared to the wild-type enzyme | Bos taurus |
W118A | site-directed mutagenesis, the mutant shows a decrease of the catalytic rate compared to the wild-type enzyme | Bos taurus |
W118A/W181A | site-directed mutagenesis, the mutant shows a decrease of the catalytic rate which is 16fold lower than the product of the effects of the two single mutations | Bos taurus |
W118F | site-directed mutagenesis, the mutant shows a decrease in activity compared to the wild-type enzyme | Bos taurus |
W118H | site-directed mutagenesis, the mutant shows a decrease in activity compared to the wild-type enzyme | Bos taurus |
W181A | site-directed mutagenesis, the mutant shows a decrease of the catalytic rate and a reduced sensitivity to nicotinamide inhibition compared to the wild-type enzyme | Bos taurus |
W181F | site-directed mutagenesis, the mutant shows a decrease in activity and an increase in ADP cyclization compared to the wild-type enzyme | Bos taurus |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
1,N6-etheno NAD+ | - |
Bos taurus | |
2'-deoxy-2'-beta-D-fluoroarabinofuranoside NAD+ | - |
Bos taurus | |
2'-deoxy-2'-beta-D-fluororibofuranoside NAD+ | non-covalent complex of the inhibitor formed with enzyme mutant E218Q, PDB ID: 3ghh, and with wild-type enzyme, PDB ID: 3kou | Bos taurus | |
3-aminopyridine | - |
Bos taurus | |
Isonicotinic acid hydrazide | - |
Bos taurus | |
nicotinamide | inhibition involves enzyme residue Trp181 | Bos taurus | |
Pyridine | non-competitive inhibition of mutant E218A by pyridine | Bos taurus |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0171 | - |
NAD+ | recombinant wild-type enzyme, pH 7.4, 37°C | Bos taurus | |
0.0175 | - |
NAD+ | recombinant mutant D147A, pH 7.4, 37°C | Bos taurus | |
0.0189 | - |
NAD+ | recombinant mutant W118A, pH 7.4, 37°C | Bos taurus | |
0.0215 | - |
NAD+ | recombinant mutant W181F, pH 7.4, 37°C | Bos taurus | |
0.0247 | - |
NAD+ | recombinant mutant E218Q, pH 7.4, 37°C | Bos taurus | |
0.0276 | - |
NAD+ | recombinant mutant E138A, pH 7.4, 37°C | Bos taurus | |
0.028 | - |
NAD+ | recombinant mutant E218A, pH 7.4, 37°C | Bos taurus | |
0.0289 | - |
NAD+ | recombinant mutant W118H, pH 7.4, 37°C | Bos taurus | |
0.0295 | - |
NAD+ | recombinant mutant E138Q, pH 7.4, 37°C | Bos taurus | |
0.0297 | - |
NAD+ | recombinant mutant W118A/W181A, pH 7.4, 37°C | Bos taurus | |
0.036 | - |
NAD+ | recombinant mutant W118F, pH 7.4, 37°C | Bos taurus | |
0.0453 | - |
NAD+ | recombinant mutant W181A, pH 7.4, 37°C | Bos taurus |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
NAD+ + H2O | Bos taurus | bovine CD38/NAD+ glycohydrolase catalyzes the hydrolysis of NAD+ to nicotinamide and ADP-ribose and the formation of cyclic ADP-ribose via a stepwise reaction mechanism | ADP-D-ribose + nicotinamide | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Bos taurus | Q9TTF5 | - |
- |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
NAD+ + H2O = ADP-D-ribose + nicotinamide + H+ | substrate binding with a crucial role of Glu218, which orients the substrate for cleavage by interacting with the N-ribosyl 2'-OH group of NAD+, stepwise ordered uni-bi kinetic mechanism, overview. Residues Trp118, Glu138, Asp147, Trp181 stabilize the ribooxocarbenium ion-like transition state mostly by electrostatic interactions | Bos taurus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
additional information | selectivity in favor of methanolysis by wild-type enzyme and mutant E138A. 1'-Azido ADP-ribose is the reaction product obtained in the presence of azide. the ADP-ribosyl cyclase activity of wild-type bCD38 isminimal | Bos taurus | ? | - |
? | |
NAD+ + H2O | bovine CD38/NAD+ glycohydrolase catalyzes the hydrolysis of NAD+ to nicotinamide and ADP-ribose and the formation of cyclic ADP-ribose via a stepwise reaction mechanism | Bos taurus | ADP-D-ribose + nicotinamide | - |
? | |
NAD+ + H2O | via a stepwise reaction mechanism | Bos taurus | ADP-D-ribose + nicotinamide | - |
? |
Synonyms | Comment | Organism |
---|---|---|
bCD38 | - |
Bos taurus |
CD38/NAD+ glycohydrolase | - |
Bos taurus |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Bos taurus |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0155 | - |
NAD+ | recombinant mutant E218A, pH 7.4, 37°C | Bos taurus | |
0.016 | - |
NAD+ | recombinant mutant W118A/W181A, pH 7.4, 37°C | Bos taurus | |
0.03 | - |
NAD+ | recombinant mutant E218Q, pH 7.4, 37°C | Bos taurus | |
0.122 | - |
NAD+ | recombinant mutant W118A, pH 7.4, 37°C | Bos taurus | |
0.49 | - |
NAD+ | recombinant mutant W181A, pH 7.4, 37°C | Bos taurus | |
0.81 | - |
NAD+ | recombinant mutant E138A, pH 7.4, 37°C | Bos taurus | |
3.5 | - |
NAD+ | recombinant mutant D147A, pH 7.4, 37°C | Bos taurus | |
6.64 | - |
NAD+ | recombinant mutant E138Q, pH 7.4, 37°C | Bos taurus | |
7.8 | - |
NAD+ | recombinant mutant W181F, pH 7.4, 37°C | Bos taurus | |
10.36 | - |
NAD+ | recombinant mutant W118F, pH 7.4, 37°C | Bos taurus | |
18.37 | - |
NAD+ | recombinant mutant W118H, pH 7.4, 37°C | Bos taurus | |
57.9 | - |
NAD+ | recombinant wild-type enzyme, pH 7.4, 37°C | Bos taurus |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.4 | - |
assay at | Bos taurus |
General Information | Comment | Organism |
---|---|---|
additional information | structure-function analysis, overview. The enzyme catalyzes the formation of beta-1'-O-methyl ADP-ribose in presence of methanol, solvolysis does not affect the overall turnover rate of NAD+ by the wild-type enzyme. Precise role of key conserved active site residues Trp118, Glu138, Asp147, Trp181 and Glu218, effects of experiments with neutral (methanol) and ionic (azide, formate) nucleophiles. Binding of 2'-fluorinated analogs of NAD+ and trappping of the reaction intermediate, detailed overview. Catalytic residue Glu138 is part of the TLEDTL signature domain, Asp147 is a highly conserved residue in the enzyme and is important for the catalytic parameters. Cooperative contribution of Trp118 and Trp181 to catalysis | Bos taurus |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.54 | - |
NAD+ | recombinant mutant W118A/W181A, pH 7.4, 37°C | Bos taurus | |
0.55 | - |
NAD+ | recombinant mutant E218A, pH 7.4, 37°C | Bos taurus | |
1.21 | - |
NAD+ | recombinant mutant E218Q, pH 7.4, 37°C | Bos taurus | |
6.4 | - |
NAD+ | recombinant mutant W118A, pH 7.4, 37°C | Bos taurus | |
10.8 | - |
NAD+ | recombinant mutant W181A, pH 7.4, 37°C | Bos taurus | |
29.3 | - |
NAD+ | recombinant mutant E138A, pH 7.4, 37°C | Bos taurus | |
200 | - |
NAD+ | recombinant mutant D147A, pH 7.4, 37°C | Bos taurus | |
220 | - |
NAD+ | recombinant mutant E138Q, pH 7.4, 37°C | Bos taurus | |
290 | - |
NAD+ | recombinant mutant W118F, pH 7.4, 37°C | Bos taurus | |
360 | - |
NAD+ | recombinant mutant W181F, pH 7.4, 37°C | Bos taurus | |
630 | - |
NAD+ | recombinant mutant W118H, pH 7.4, 37°C | Bos taurus | |
3380 | - |
NAD+ | recombinant wild-type enzyme, pH 7.4, 37°C | Bos taurus |