Part:BBa_K3629009
Trichoderma reesei EGII with FLAG tag
Endoglucanase II coding sequence from Trichoderma reesei with FLAG tag.
Usage and Biology
Yarrowia lipolytica is an emerging chassis in the molecular biology community. Its unique metabolic properties and efficient protein production and secretion mechanisms make it a desirable chassis for heterologous protein expression/secretion. In fact, it has been shown to have better secretory mechanisms than Saccharomyces cerevisiae (1). Therefore, using this chassis to secrete cellulase enzymes- which are enzymes that require high levels of secretion, is well suited.
Fully functional cellulase is composed of:
- Endoglucanases (EG) which randomly cleave internal beta-bonds of cellulose polymers to make them shorter
- Cellobiohydrolases (CBH or exoglucanases) which cleave the shorter polymers to make cellobiose
- CBHI= Acts on reducing end of sugar molecule
- CBHII= Acts on non-reducing end of sugar molecule
- Beta-glucosidases (BGS) which cleave the cellobiose disaccharide to free glucose units
These proteins must be in the correct proportions to each other to efficiently degrade cellulose.
EGs specifically are the most expressed cellulases in Trichoderma reesei while EGI has the most efficient hydrolysis on crystalline cellulose (2) The activity of endoglucanases is important to produce more exposed ends for exoglucanase or cellobiohydrolase activity.
Design
The native signal peptide from T. reesei was removed so it would not interfere with fused secretion tags native to Y. lipolytica
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 283
Illegal NgoMIV site found at 817
Illegal NgoMIV site found at 1135 - 1000COMPATIBLE WITH RFC[1000]
References
//chassis/eukaryote/yeast
None |