Purification and characterization of a glutaminase enzyme accounting for the majority of glutaminase activity in Aspergillus sojae under solid-state culture
| Autor: | Kotaro Ito, Yoshiki Hanya, Yasuji Koyama |
|---|---|
| Rok vydání: | 2012 |
| Předmět: |
Molecular Sequence Data
Biology Aspergillus sojae Applied Microbiology and Biotechnology Glutaminase activity Substrate Specificity Aspergillus oryzae Glutaminase Enzyme Stability DNA Fungal Phylogeny chemistry.chemical_classification Aspergillus Molecular mass Sequence Homology Amino Acid Fungal genetics Temperature General Medicine Sequence Analysis DNA Hydrogen-Ion Concentration biology.organism_classification Molecular biology Molecular Weight Enzyme chemistry Biochemistry Chromatography Gel Electrophoresis Polyacrylamide Gel Protein Multimerization Gene Deletion Biotechnology |
| Zdroj: | Applied microbiology and biotechnology. 97(19) |
| ISSN: | 1432-0614 |
| Popis: | Glutaminase, an enzyme that hydrolyzes l-glutamine to l-glutamate, plays an important role in the production of fermented foods by enhancing the umami taste. In this study, we found ten glutaminase genes in the Aspergillus sojae genome by conducting a BLAST search of the characterized glutaminase sequence. We subsequently constructed glutaminase gene disruptants. The glutaminase activity of the gahB disruptant was decreased by approximately 90 % in A. sojae and Aspergillus oryzae, indicating that this enzyme (GahB) accounted for the majority of the glutaminase activity in Aspergillus species. Subsequently, GahB protein was purified from the AsgahB-overexpressing transformant and characterized. The molecular mass was estimated to be approximately 110 and 259 kDa by SDS-PAGE and gel filtration chromatography, respectively, indicating that the native form of AsGahB was a dimer. The optimal pH was 9.0, and the optimal temperature was 50 °C. Analysis of substrate specificity revealed that AsGahB had peptidoglutaminase-asparaginase activity, similar to AsGahA, but preferred free l-glutamine to free l-asparagine, C-terminal glutaminyl, and asparaginyl residues in peptides. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full text from SpringerLink