Development of fluorescent labeling method for AHL receptors responding to cross-kingdom signaling

2022 Fiscal Year Final Research Report

• Project/Area Number: 21K14765
• Research Category: Grant-in-Aid for Early-Career Scientists
• Allocation Type: Multi-year Fund
• Review Section: Basic Section 38020:Applied microbiology-related
• Research Institution: Utsunomiya University
• Principal Investigator: Nasuno Eri 宇都宮大学, 工学部, 助教 (80709329)
• Project Period (FY): 2021-04-01 – 2023-03-31
• Keywords: クオラムセンシング / N-アシルホモセリンラクトン / LuxR familyタンパク質 / 蛍光プローブ

Outline of Final Research Achievements

Some Gram-negative bacteria have a quorum sensing (QS) that controls the expression of genes, depending on the intracellular concentration of an acyl homoserine lactone (AHL) as a signal molecule. In this study, we developed a unique fluorescent probe to specifically label the LuxR which is known as an AHL receptor. 4-nitrobenzo-2-oxa-1,3-diazole (NBD) groups were introduced to the acyl chain end of AHLs with different chain length as a chromophore. CviR derived from Chromobacterium violaceum, which regulates to produce a purple pigment in response to AHL, was selected as a model LuxR. As a result of the fluorescence labeling test, all AHL-NBD probes with acyl chain lengths from C6 to C12 could be fluorescently labeled the LuxR, especially the long-chain AHL-NBD probe showed particularly high fluorescence intensity. Since the AHL-NBD probes were able to label the proteins both in vitro and in vivo, we will further evaluate the labeling activity of our probes for the unknown LuxR.

Free Research Field

生物工学および微生物生態学

Academic Significance and Societal Importance of the Research Achievements

データベースに登録されている3000種以上の細菌ゲノムから同定されたAHL受容体の約80%は、ペアになるAHL合成酵素が存在しない。AHL受容体を特異的に蛍光標識可能なAHL-NBDプローブは、これらのAHL受容体のみを保有する細菌の細胞間情報伝達機構を簡便に特定する新規解析ツールと言える。このAHL-NBDプローブを用いることで細菌にとどまらず細菌と競争・共生関係を構築している動植物まで含む広範な生物種を対象とした細胞間情報伝達機構による遺伝子発現制御システムの解明に繋がると期待される。