Near Ultraviolet Absorption Arising from Lysine Residues in Close Proximity: A Probe to Monitor Protein Unfolding and Aggregation in Lysine-Rich Proteins

Abstract

There is a need for an intrinsic spectral probe to monitor key events like protein unfolding and aggregation in a rapid and unambiguous manner. Protein aggregation is an important issue, but ironically there is a dearth of simple techniques to directly detect the presence of aggregates in solution. We report here the hitherto undiscovered electronic absorption around 300-350 nm in aqueous solutions (pH 7) of human serum albumin (HA), calf thymus histone, and poly-L-lysine. The above spectra were significantly absent in controls like subtilisin carlsberg, mutant barstar, and lysozyme. The possibilities that Rayleigh scattering or impurities could account for the above spectra were checked and ruled out. Based on the analysis of available three-dimensional structures from PDB and our earlier work on the lysine amino acid, an intramolecular interaction between lysine side chains in close spatial proximity was deduced to be the origin for the above spectra. The utility of Lys-Lys interaction in detecting protein unfolding and aggregation in a lysine-rich protein like calf thymus histone using near-ultraviolet absorption is demonstrated.

Weak Intrinsic Luminescence in Monomeric Proteins Arising from Charge Recombination

“…Several years ago, our group had reported electronic absorption and subsequent luminescence originating from dense aqueous solutions of l -Lysine·HCl . This was later corroborated by other groups as well. − Similar absorption was also evident in dilute aqueous solutions of lysine-rich proteins, like human serum albumin (HuSA), poly- l -Lysine·HCl, and calf thymus histone . Luminescence identical to lysine solutions was also observed in dilute solutions of HuSA protein, which is rich in charged amino acids like Lys and Glu (Homchaudhuri and Swaminathan, unpublished).…”

“…The higher molar absorptivity in proteins in comparison with free amino acids arises from the large effective concentration of different charged amino acids lying in close proximity to each other in a protein owing to the 3D fold, unlike free amino acids that are not connected by peptide bonds. This was postulated long ago by our group. , Thus, the slope magnitude and therefore the luminescence sensitivity reflects the richness of spatial interactions among charged amino acids in the protein.…”

“…This was postulated long ago by our group. 11,13 Thus, the slope magnitude and therefore the luminescence sensitivity reflects the richness of spatial interactions among charged amino acids in the protein.…”

“…8−10 Similar absorption was also evident in dilute aqueous solutions of lysine-rich proteins, like human serum albumin (HuSA), poly-L-Lysine• HCl, and calf thymus histone. 11 Luminescence identical to lysine solutions was also observed in dilute solutions of HuSA protein, which is rich in charged amino acids like Lys and Glu (Homchaudhuri and Swaminathan, unpublished) headgroup, the chromophore giving rise to observed luminescence was not obvious.…”

A Comprehensive Review of Computation-Based Metal-Binding Prediction Approaches at the Residue Level

“…Amino acid pairs, or dipeptides, are based on the observation that amino acid pairs show different propensities in protein structure and function. For instance, pairs of lysine are found present in close spatial vicinity [ 67 ]. Moreover, the concept of k -spaced amino acid pairs is introduced in [ 68 ].…”