Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli as a His-tagged fusion protein | Microbacterium sp. |
Protein Variants | Comment | Organism |
---|---|---|
A264G | relative hydrolytic activity: 20.4% compared to levanbiohydrolase from Microbacterium laevaniformans. Relative hydrolytic activity of wild-type: 10.7% | Microbacterium sp. |
additional information | residues that are critical to the intramolecular fructosyl transfer reaction of LFTase are analysed. An error-prone PCR mutagenesis process is carried out, and the mutants that led to a shift in activity from transfructosylation towards hydrolysis of levan are screened by the dinitrosalicylic acid (DNS) method. Selected mutants reveal two major products: one is a di-D-fructose-2,6':6,2'-dianhydride (DFAIV) and the other is a levanbiose. The detected levanbiose corresponds to the reaction product from LFTase lacking transferring activity N85S substitution is responsible for the strong enhancement of the hydrolytic activity | Microbacterium sp. |
N238K | relative hydrolytic activity: 23.6% compared to levanbiohydrolase from Microbacterium laevaniformans. Relative hydrolytic activity of wild-type: 10.7% | Microbacterium sp. |
N85S | relative hydrolytic activity: 34.8% compared to levanbiohydrolase from Microbacterium laevaniformans. Relative hydrolytic activity of wild-type: 10.7% | Microbacterium sp. |
N85S/A264G | relative hydrolytic activity: 49.9% compared to levanbiohydrolase from Microbacterium laevaniformans. Relative hydrolytic activity of wild-type: 10.7% | Microbacterium sp. |
N85S/D268E | relative hydrolytic activity: 30% compared to levanbiohydrolase from Microbacterium laevaniformans. Relative hydrolytic activity of wild-type: 10.7% | Microbacterium sp. |
N85S/N224K/Q229E/T253N | relative hydrolytic activity: 37% compared to levanbiohydrolase from Microbacterium laevaniformans. Relative hydrolytic activity of wild-type: 10.7% | Microbacterium sp. |
P313R/F518L | relative hydrolytic activity: 15.8% compared to levanbiohydrolase from Microbacterium laevaniformans. Relative hydrolytic activity of wild-type: 10.7% | Microbacterium sp. |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Microbacterium sp. | - |
- |
- |
Purification (Comment) | Organism |
---|---|
after Ni-NTA chromatogrphy the enzyme solution is further purified on a Hi-Prep 16/60 Sephacryl S-200 high resolution column using fast-performance liquid chromatography | Microbacterium sp. |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
levan | - |
Microbacterium sp. | 5 di-D-fructose-2,6':2',6 dianhydride + ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
levan fructotransferase | - |
Microbacterium sp. |
LFTase | - |
Microbacterium sp. |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Microbacterium sp. |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7 | - |
assay at | Microbacterium sp. |