LiLac - a biosensor for Lactate

LiLac is a biosensor for lactate with a fluorescence-lifetime readout ^[1]. The fluorescent protein in LiLac is mTurquoise, a low pKa fluorescent protein commonly used in lifetime-contrast sensors. The ligand-binding protein in LiLac is the lactate-binding Cache domain of TlpC. Cache-domain-containing proteins sense extracellular nutrients to guide chemotaxis in bacteria; the N- and C- termini of the nutrient-sensing regions in Cache proteins (like the lactate-binding region of TlpC) are close in space, facilitating modular design. The combination of this modularity and the large diversity of ligands that Cache domains can sense have made this domain family an attractive starting point for building sensors. The mTurquoise is split in the middle of its seventh β-strand (β7) and then inserted the extracellular domain of TlpC at this position ^[2]. By engineering the connections between split mTurquoise and TlpC using a microfluidics-based screen, we developed LiLac, a lactate biosensor with a robust ~1.2 ns decrease (−35%) in fluorescence lifetime as lactate binds.

LiLac has the following parts - , an N terminal domain, the lactate binding , a and .

The current report describes the structural aspect of LiLac biosensor (as a part of the course BI3323-Aug2025), and this is entirely adapted from the work entitled " State-dependent motion of a genetically encoded fluorescent biosensor” by Rosena et al ^[3].

Mechanism of action

When lactate binds, the fluorescent protein rotates ~150° relative to the TlpC domain and translates by 30 Å as shown in the figure that depicts the twist upon lactate binding.

The first contributing feature is a “lid” that folds on top of lactate. The tip of this loop portion of LiLac in the absence of lactate, but it moves ~11 Å in the binding pocket.

Second, a at the back of the binding pocket, near its very C-terminus, shortens by one amino acid to accommodate lactate when it binds. This shortening is associated with a retraction of the TlpC “tail” in LiLac, which protrudes in the lactate-free form.

Disease

Relevance

Structural highlights

This is a sample scene created with SAT to by Group, and another to make of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.