Intrinsic fluorescence in peptides
Since the mid-20th century, the consensus has been that the fluorescence of proteins is derived exclusively from the presence of aromatic groups, more specifically the amino acids tryptophan, tyrosine, and phenylalanine. This has largely dictated how researchers interpret the photophysical properties of biological matter. However, this aromatic-only rule was preceded by much earlier observations that hinted at a more complex reality. As early as 1928, researchers like P. Wels reported a mysterious blue fluorescence in serum albumin that could not be explained by the presence of traditional chromophores. Over the years, similar reports of non-aromatic or intrinsic photoluminescence appeared in various systems, including starch and synthetic polymers, yet the phenomenon remained largely unexplored due to a messy theoretical landscape and a lack of systematic studies.
In recent years, the study of Intrinsic Fluorescence (IF) has resurfaced as a significant fundamental challenge that defies our conventional views of photoluminescence. Historically, research was hindered by the assumption that IF only occurred in the aggregate state, such as in amyloid fibers or large protein clusters. Our research directly challenges this asumption by focusing on rationally designed zwitterionic α-helical peptides and miniproteins that exhibit intense IF in solution. By using these structurally well-defined scaffolds, we are uncovering the specific sequence and structure relationships that allow non-aromatic polypeptides to glow. This work is not only redefining the concept of the fluorophore but also laying the foundation for a new enabling technology: small, genetically encodable IF tags that could serve as powerful imaging tools for biomedical research.
Non-Aromatic Fluorescence in alpha-Helical Peptides
Our systematic analysis, recently published in Cell Reports Physical Science, provides the first evidence that NAF can be engineered into simple, molecular peptides in solution. We demonstrated that zwitterionic single α-helices (SAHs), composed exclusively of non-aromatic glutamic acid and lysine or arginine residues, are UV-active and exhibit distinct blue fluorescence. The study reveals that this emission is intimately linked to the α-helical fold; when the peptide adopts its structured conformation, the charges are clustered into a network of salt bridges that facilitates through-space electronic interactions.
We further showed that conservative mutations, such as replacing lysine with ornithine or arginine, significantly modulate the intensity and wavelength of the emission. Beyond its fundamental interest, we exploited this IF phenomenon to develop a new sensing mechanism: using the helical peptide as an antenna to sensitize the luminescence of lanthanide ions like Terbium (III) and Europium (III). These findings prove that IF is a real, programmable property of soluble polypeptides, opening the door to the development of genetically encodable fluorescent markers that do not rely on traditional aromatic chemistry.
