Molecular and enzymatic characterization of a levan fructotransferase from Microbacterium sp. AL-210
| Autor: | Jaeho Cha, Tae Ho Lee, Sung-Jae Yang, Na Hee Park |
|---|---|
| Rok vydání: | 2001 |
| Předmět: |
Levan fructotransferase
Molecular Sequence Data Size-exclusion chromatography Microbacterium Bioengineering Applied Microbiology and Biotechnology Actinomycetales Amino Acid Sequence Cloning Molecular Peptide sequence chemistry.chemical_classification Chromatography Base Sequence biology Chemistry Nucleic acid sequence Sequence Analysis DNA General Medicine biology.organism_classification Enzyme assay Enzyme Isoelectric point Hexosyltransferases Biochemistry biology.protein Biotechnology |
| Zdroj: | Journal of Biotechnology. 91:49-61 |
| ISSN: | 0168-1656 |
| DOI: | 10.1016/s0168-1656(01)00288-7 |
| Popis: | Microbacterium sp. AL-210 producing a novel levan fructotransferase (LFTase) was screened from soil samples. The LFTase was purified to homogeneity by (NH4)2SO4 fractionation, column chromatography on Resource Q, and Superdex 200HR. The molecular weight of the purified enzyme was estimated to be approximately 46 kDa by both SDS-PAGE and gel filtration, and the enzyme's isoelectric point was pH 4.8. The major product produced from the levan hydrolysis by the enzyme reaction was identified by atmospheric pressure ionization mass spectrometry and NMR analysis as di-D-fructose-2,6':6,2'-dianhydride (DFA IV). The optimum pH and temperature for DFA IV production were 7.0 and 40 degrees C, respectively. The enzyme was stable at a pH range 7.0-8.0 and up to 40 degrees C. The enzyme activity was inhibited by FeCl2 and AgNO3. The enzyme converted the levan to DFA IV, with a conversion yield of approximately 44%. A gene encoding the LFTase (lftM) from Microbacterium sp. AL-210 was cloned and sequenced. The nucleotide sequence included an ORF of 1593 nucleotides, which is translated into a protein of 530 amino acid residues. The predicted amino acid sequence of the enzyme shared 79% of the identity and 86% of the homology with that of Arthrobacter nicotinovorans GS-9. |
| Databáze: | OpenAIRE |
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