Structural and functional analysis of novel cold-adapted carbohydrate hydrolases from Eisenia fetida and application of their enzymes

2014 Fiscal Year Final Research Report

• Project/Area Number: 24580481
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Boundary agriculture
• Research Institution: Osaka Prefecture University
• Principal Investigator: UEDA mitsuhiro 大阪府立大学, 生命環境科学研究科(系), 准教授 (50254438)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Keywords: 糖質分解酵素 / 低温適応性 / ミミズ / キチナーゼ / アミラーゼ / X線結晶構造解析 / セルラーゼ

Outline of Final Research Achievements

The present study identified a cold adapted cellulase named endo-1,4-beta-glucanase (EF-EG2) from the earthworm Eisenia fetida. The gene was cloned in the cold-shock expression vector and functionally expressed in Escherichia coli. The gene consists of 1368 bp encoding 456 amino acid residues. Purified recombinant EF-EG2 hydrolyzed soluble cellulose (carboxymethyl cellulose), but not insoluble (powdered cellulose) or crystalline (Avicel) cellulose substrates. The enzyme exhibited significant activity at 10oC (38% of the activity at optimal 40oC) and was stable at pH 5.0~9.0, with an optimum pH of 5.5. This new cold-adapted cellulase could potentially improve the cost effectiveness of biofuel production. The overall structure of EF-EG2 has an (alpha/alpha)6 barrel fold which contains a putative activesite cleft and a negatively charged surface. This structural information helps us understand the catalytic and cold adaptation mechanisms of EF-EG2.

Free Research Field

生物資源利用学