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Literature summary for 1.13.12.7 extracted from
- SpyTag/SpyCatcher cyclization enhances the thermostability of firefly luciferase (2016), PLoS ONE, 11, e0162318 .
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| recombinant coexpression of the luciferase with fused cyclization proteins SpyCatcher at the N-terminus and SpyTag at the C-terminus (pyCatcher-LUC-SpyTag) in Escherichia coli strain BL21(DE3) | Photinus pyralis |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| additional information | firefly luciferase from Photinus pyralis is cyclized in vivo by fusing SpyCatcher at the N-terminus and SpyTag at the C-terminus. SpyTag can spontaneously form a covalent isopeptide bond with its protein partner Spy-Catcher. Circular LUC is more thermostable at 25-55°C and alkali-tolerant than the wild-type, without compromising the specific activity. Preparation of an N-terminally and C-terminally shortened form of the SpyCatcher protein and enzyme cyclization using this truncated form leads to even more thermostability of the construct than the full-length SpyCatcher form, method optimization | Photinus pyralis |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| D-firefly luciferin + O2 + ATP | Photinus pyralis | - |
firefly oxyluciferin + CO2 + AMP + diphosphate + hv | - |
? |  |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Photinus pyralis | P08659 | - |
- |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| D-firefly luciferin + O2 + ATP | - |
Photinus pyralis | firefly oxyluciferin + CO2 + AMP + diphosphate + hv | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| ? | x * 61000, recombinant wild-type enzyme, SDS-PAGE, x * 80000, recombinant cyclized enzyme SpyCatcher-LUC-SpyTag, SDS-PAGE | Photinus pyralis |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| firefly luciferase | - |
Photinus pyralis |
| Luc | - |
Photinus pyralis |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 25 | - |
wild-type enzyme | Photinus pyralis |
| 35 | - |
recombinant circular enzyme | Photinus pyralis |
Temperature Range [°C]
| Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 10 | 60 | temperature activity range, recombinant wild-type enzyme and recombinant circular enzyme | Photinus pyralis |
Temperature Stability [°C]
| Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 16.7 | - |
Tm value of recombinant wild-type LUC | Photinus pyralis |
| 37.4 | - |
T50 for recombinant wild-type LUC | Photinus pyralis |
| 40.3 | - |
T1/2 value of recombinant cyclized LUC | Photinus pyralis |
| 47.5 | - |
T50 for recombinant cyclized LUC | Photinus pyralis |
| 49.2 | - |
T50 value of recombinant cyclized LUC with truncated Spy-Catcher cyclization protein | Photinus pyralis |
| 51.4 | - |
T1/2 value of recombinant cyclized LUC with truncated Spy-Catcher cyclization protein | Photinus pyralis |
| 53.5 | - |
Tm value of recombinant wild-type LUC | Photinus pyralis |
| 70.2 | - |
Tm value of recombinant cyclized LUC | Photinus pyralis |
| 73.1 | - |
Tm value of recombinant cyclized LUC with truncated Spy-Catcher cyclization protein | Photinus pyralis |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 8 | - |
wild-type enzyme | Photinus pyralis |
| 9 | - |
recombinant circular enzyme | Photinus pyralis |
pH Range
| pH Minimum | pH Maximum | Comment | Organism |
|---|---|---|---|
| 2.5 | 10.5 | pH activity range, recombinant circular enzyme | Photinus pyralis |
| 4 | 10 | pH activity range, recombinant wild-type enzyme | Photinus pyralis |
pH Stability
| pH Stability | pH Stability Maximum | Comment | Organism |
|---|---|---|---|
| 8 | 10 | recombinant circular LUC is stable at a pH range of 8.0-10.0, maintaining nearly 100% activity after incubation in various pH buffers for 30 min. Recombinant wild-type LUC maintains only 80% of its original activity after incubating at pH 10.0 for 30 min | Photinus pyralis |
Cofactor
| Cofactor | Comment | Organism | Structure |
|---|---|---|---|
| ATP | - |
Photinus pyralis | |
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