Cloned (Comment) | Organism |
---|---|
expression of wild-type and of R42K, R42L and G41R/R42L mutant enzymes in Escherichia coli JM109 as His6-tagged proteins | Escherichia coli |
Crystallization (Comment) | Organism |
---|---|
- |
Escherichia coli |
Protein Variants | Comment | Organism |
---|---|---|
G41D | mutant enzyme with decreased activity | Escherichia coli |
G41K | mutant enzyme with decreased activity | Escherichia coli |
G41L | mutant enzyme with decreased activity | Escherichia coli |
G41R | G41R/R42L double mutant with low activity, G41R: decreased activity | Escherichia coli |
R38K | mutant enzyme with 2.2fold increased Km, no effect on kcat | Escherichia coli |
R38L | mutant enzyme with up to 12fold lower specific activity, 6fold increased Km and 5fold decreased kcat | Escherichia coli |
R42K | mutant enzyme with lower activity, 16fold increased Km | Escherichia coli |
R42L | single mutant and G41R/R42L double mutant with low activity, R42L: 17fold increased Km and 79fold decreased kcat | Escherichia coli |
R42Q | mutant enzyme with lower activity, 16-17fold increased Km | Escherichia coli |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | values for wild-type and mutant enzyme with wild-type and mutant initiator tRNAs | Escherichia coli | |
0.00054 | - |
L-methionyl-tRNAfMet | wild-type enzyme | Escherichia coli | |
0.00066 | - |
L-methionyl-tRNAfMet | G41R mutant enzyme | Escherichia coli | |
0.00102 | - |
L-methionyl-tRNAfMet | G41K mutant enzyme | Escherichia coli | |
0.00121 | - |
L-methionyl-tRNAfMet | R38K mutant enzyme | Escherichia coli | |
0.00162 | - |
L-methionyl-tRNAfMet | G41L mutant enzyme | Escherichia coli | |
0.003 | - |
L-methionyl-tRNAfMet | G41D mutant enzyme | Escherichia coli | |
0.00324 | - |
L-methionyl-tRNAfMet | R38L mutant enzyme | Escherichia coli | |
0.00834 | - |
L-methionyl-tRNAfMet | G41R/R42L double mutant enzyme | Escherichia coli | |
0.00866 | - |
L-methionyl-tRNAfMet | R42K mutant enzyme | Escherichia coli | |
0.009 | - |
L-methionyl-tRNAfMet | R42L mutant enzyme | Escherichia coli | |
0.0096 | - |
L-methionyl-tRNAfMet | R42Q mutant enzyme | Escherichia coli |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
10-formyltetrahydrofolate + L-methionyl-tRNAfMet + H2O | Escherichia coli | involved in initiation of protein biosynthesis | tetrahydrofolate + N-formylmethionyl-tRNAfMet | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Escherichia coli | - |
- |
- |
Purification (Comment) | Organism |
---|---|
purification of wild-type and of R42K, R42L and G41R/R42L mutant recombinant enzymes, expressed in Escherichia coli JM109 | Escherichia coli |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
additional information | - |
- |
Escherichia coli |
Storage Stability | Organism |
---|---|
-20°C, 20 mM imidazole, 10 mM 2-mercaptoethanol, 50% glycerol, 150 mM KCl, stable | Escherichia coli |
-20°C, 20 mM imidazole, pH 7.5, 10 mM 2-mercaptoethanol, 50% glycerol, 150 mM KCl, 2-4 months, stable | Escherichia coli |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
10-formyltetrahydrofolate + L-methionyl-tRNAfMet | - |
Escherichia coli | tetrahydrofolate + N-formylmethionyl-tRNAfMet | - |
r | |
10-formyltetrahydrofolate + L-methionyl-tRNAfMet + H2O | specificity determinants at the end of the acceptor stem of tRNAfMet | Escherichia coli | tetrahydrofolate + N-formylmethionyl-tRNAfMet | - |
? | |
10-formyltetrahydrofolate + L-methionyl-tRNAfMet + H2O | invariant Arg-42 within the 16-amino acid insertion loop, residues 34-49, plays a very important role both in steps of substrate binding and in catalysis, Arg-42 makes functional contacts with the determinants in the acceptor stem of the initiator tRNA | Escherichia coli | tetrahydrofolate + N-formylmethionyl-tRNAfMet | - |
? | |
10-formyltetrahydrofolate + L-methionyl-tRNAfMet + H2O | specificity is restricted to methionyl-tRNAfMet | Escherichia coli | tetrahydrofolate + N-formylmethionyl-tRNAfMet | - |
? | |
10-formyltetrahydrofolate + L-methionyl-tRNAfMet + H2O | important role of a 16-amino acid insertion loop in enzyme for tRNA recognition | Escherichia coli | tetrahydrofolate + N-formylmethionyl-tRNAfMet | - |
? | |
10-formyltetrahydrofolate + L-methionyl-tRNAfMet + H2O | involved in initiation of protein biosynthesis | Escherichia coli | tetrahydrofolate + N-formylmethionyl-tRNAfMet | - |
? | |
additional information | enzyme structure | Escherichia coli | ? | - |
? |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Escherichia coli |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | values for wild-type and mutant enzyme with wild-type and mutant initiator tRNAs | Escherichia coli | |
0.45 | - |
L-methionyl-tRNAfMet | G41R/R42L double mutant enzyme | Escherichia coli | |
0.483 | - |
L-methionyl-tRNAfMet | R42L mutant enzyme | Escherichia coli | |
3.1 | - |
L-methionyl-tRNAfMet | R42Q mutant enzyme | Escherichia coli | |
8.27 | - |
L-methionyl-tRNAfMet | R38L mutant enzyme | Escherichia coli | |
11.4 | - |
L-methionyl-tRNAfMet | G41R mutant enzyme | Escherichia coli | |
14.6 | - |
L-methionyl-tRNAfMet | G41K mutant enzyme | Escherichia coli | |
27.7 | - |
L-methionyl-tRNAfMet | R42K mutant enzyme | Escherichia coli | |
31.5 | - |
L-methionyl-tRNAfMet | G41L mutant enzyme | Escherichia coli | |
33 | - |
L-methionyl-tRNAfMet | G41D mutant enzyme | Escherichia coli | |
39.1 | - |
L-methionyl-tRNAfMet | wild-type enzyme | Escherichia coli | |
46.3 | - |
L-methionyl-tRNAfMet | R38K mutant enzyme | Escherichia coli |