Investigation of the enzymatic reaction of vacuolar-type ATPase by isolation and characterization of mutants of bacterial sodium-translocating V-ATPase

2016 Fiscal Year Final Research Report

• Project/Area Number: 26850234
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Applied molecular and cellular biology
• Research Institution: Ehime University
• Principal Investigator: Kawada Miyuki 愛媛大学, 農学研究科, 准教授 (10454498)
• Research Collaborator: Kakinuma Yoshimi
• Project Period (FY): 2014-04-01 – 2017-03-31
• Keywords: V-ATPase / ナトリウム輸送性 / 腸球菌

Outline of Final Research Achievements

Vacuolar-type ATPases (V-ATPases) structurally resemble F-ATPases and function as proton pump in acidic organelles and plasma membranes of eukaryotic cells as well as in bacteria. The Enterococcus hirae V-ATPase is a unique variant of V-ATPase, which recognizes sodium and lithium with near equal affinity and is indispensable for salt tolerance of this bacterium at alkaline pH. We previously indicated that NtpK(E139D) mutant strain lost tolerance to sodium but not to lithium, and suggested that the E139 residue is indispensable for the enzymatic activity of E. hirae V-ATPase. In an attempt to isolate the suppressor mutations for NtpK(E139D), we isolated two mutants, NtpK(E50K/E139D) and NtpK(V138I/E139D). Interestingly, the single point mutation of NtpK(E50K) or NtpK(E50D) decreased the activities of V-ATPase to approximately 50% of the wild-type, suggesting that E50 is one of the important residues for the enzymatic activity of E. hirae V-ATPase.

Free Research Field

生化学