Functional analysis of novel bacterial glycosyltransferases

2017 Fiscal Year Final Research Report

• Project/Area Number: 15K06976
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: Kyushu University
• Principal Investigator: OKINO NOZOMU 九州大学, 農学研究院, 准教授 (90363324)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Keywords: 糖脂質 / 緑膿菌 / 糖転移酵素 / グルクロノシルジアシルセラミド / グルクロノシルジアシルグリセロール

Outline of Final Research Achievements

In this study, we aimed to elucidate the physiological functions of bacterial glycolipids by functional analysis of ceramide glucuronosyltransferase (Cer GlcAT) of Zymomonas mobilis and diacylglycerol glucuronosyltransferase (DAG GlcAT) of Pseudomonas aeruginosa.
We first cloned and expressed Z. mobilis Cer GlcAT in E. coli and purified the enzyme using Ni-affinity and gel filtration chromatography. Using the purified enzyme, we revealed that the enzyme was highly specific for UDP-glucuronic acid and ceramide as donor and acceptor substrates, respectively. We also established the knockout strain and elucidated that this enzyme was the sole Cer GlcAT of Z. mobilis. Next, we cloned PA0842 of P. aeruginonsa and expressed it in E. coli. We detected the DAG GlcAT activity when UDP-glucuronic acid and diacylglycerol were used as donor and acceptor substrates, respectively. Furthermore, we found that PA0842 was induced to produce glucuronosyl DAG under phosphate-deficient conditions.

Free Research Field

生物化学