High-performance genetically-encoded green and red fluorescent biosensors for pyruvate

Shosei Imai; Saaya Hario; Carl Suerte; Issei Yamaguchi; Kenryo Sakoi; Adam Thuen; Brekken Meznarich; Mikhail A. Drobizhev; Takuya Terai; Kei Takahashi; Robert E. Campbell

doi:10.1101/2025.04.17.649293

Molecular BiologybioRxivApr 17, 2025

Preprint

High-performance genetically-encoded green and red fluorescent biosensors for pyruvate

Shosei Imai iD, Saaya Hario iD, Carl Suerte, Issei Yamaguchi, Kenryo Sakoi, Adam Thuen iD, Brekken Meznarich, Mikhail A. Drobizhev iD, Takuya Terai iD, Kei Takahashi iD, Robert E. Campbell iD

PDF DOI

Post to X

Threads on X Post to Bluesky Threads on Bluesky

Abstract

Pyruvate is the end-product of glycolysis and a central metabolite involved in many biochemical pathways. However, a lack of high-performance (i.e., {Delta}F/F0 > 10) single fluorescent protein (FP)-based biosensors has hindered efforts to investigate the physiological role of pyruvate. Here, we present the GreenPy1 and ApplePy1 series, which are green FP (GFP)-based and red FP (RFP)-based pyruvate biosensors, respectively. Both series exhibit large fluorescence intensity change ({Delta}F/F0 [~] 20 to >40) and a range of affinities (10s of M to several mM). We demonstrate the utility of these pyruvate biosensors for multicolor imaging of metabolite concentration changes in mammalian cells.