Construction of a novel cellulolysis system utilizing Tetrahymena

2017 Fiscal Year Final Research Report

• Project/Area Number: 15K12263
• Research Category: Grant-in-Aid for Challenging Exploratory Research
• Allocation Type: Multi-year Fund
• Research Field: Design and evaluation of sustainable and environmental conscious system
• Research Institution: Kanazawa University
• Principal Investigator: Endoh Hiroshi 金沢大学, 自然システム学系, 准教授 (20272932)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Keywords: テトラヒメナ / 分泌ベクター / シロアリ腸内共生ベン毛虫 / エンドグルカナーゼ / セロビオヒドラーゼ

Outline of Final Research Achievements

Construction of a novel transformation vector for expressing and secreting foreign cellulase genes in the ciliate Tetrahymena thermopile. This vector consists of 3 separate parts: 1) a cassette carrying drug resistant gene for screening after bombardment, 2) an expression cassette with a secretion signal derived from Tetrahymena cystein protease 4 gene, 3) flanking sequences at both sides of beta-tubulin gene for homologous recombination. Introducing 4 different genes (endoglucanase (EG) 5 and 7, cellobiohydrolase (CBH) and lignin esterase Cip2-like) from the termite symbiotic protists for cellulolysis into the vector, their activities were measured after induction of gene expression and subsequent secretion outside cells, respectively. EG5 and 7 were efficiently expressed, showing strong activity. Lignin esterase Cip2-like activity was slightly detected. No CBH activity has been confirmed, suggesting difficult folding due to the complicated protein structure.

Free Research Field

進化原生生物学