Quantifying amino acid and protein substitution using Raman spectroscopy

Abstract

Raman spectroscopy can be a powerful tool for the characterization of modified amino acids and proteins. In addition to the potential for quantitative results, it offers the advantage of not requiring any sample preparation. Modification of amines and thiols on amino acids and proteins are common reactions used for medical, biological, food, and agricultural purposes. We hypothesized that the Raman spectrum could be used to quantify the reactions and would be more informative than typical characterization techniques such as the ninhydrin test. To prove the hypothesis, the amino acids alanine, cysteine, and lysine were modified with ethyl vinyl sulfone (EVS) using a nucleophilic addition reaction known as the Michael addition and the product was characterized using Raman spectroscopy. The Raman spectroscopy results were compared with ultraviolet-visible spectroscopy results based on ninhydrin analysis of the modified amino acids. The Raman spectroscopy analysis was able to discern site-specific reactions on the amino acids and suggested that more amino acid moieties were substituted than predicted using the ninhydrin test alone. Substitution of the full protein ovalbumin with EVS showed similar results. The ninhydrin test showed the substitution of primary amines and thiols but could not detect substitution of secondary amines remaining after loss of the primary amine.

Glycerol citrate polyesters produced through heating without catalysis

“…Numerous issues such as price uncertainty, manufacturing costs, environmental disposal, and geo‐political concerns are affecting the petroleum‐based polymer industry. As an alternative, since 1980s there is an increasing interest in the development of biodegradable nonpetroleum‐based polymers 1–4. However, the price of these nonpetroleum‐based polymers [e.g., poly(lactic acid), PLA] is high and not competitive with petroleum‐based polymers (e.g., polyethylene terephthalate, PET) 4.…”

“…As an alternative, since 1980s there is an increasing interest in the development of biodegradable nonpetroleum‐based polymers 1–4. However, the price of these nonpetroleum‐based polymers [e.g., poly(lactic acid), PLA] is high and not competitive with petroleum‐based polymers (e.g., polyethylene terephthalate, PET) 4. Therefore, the development of lower cost nonpetroleum‐based polymers is needed.…”

Self-Assembling Functionalized Amino Acids into Unusual Shapes

“…22 It was found that lysine preferably substitutes at the α-carbon at pH 9, which would be expected from the pK a of the α-and ε-amine. Using FT-IR spectroscopy, substitution reactions can be followed in solution through the change in the amine on the amino acid (Fig.…”

Influencing factors of external fluorescence seeding enhancing stimulated Raman scattering in liquid‐core optical fiber

“…This experimental result can be explained by theory of SRS enhancement by fluorescence. The fluorescent seeds can linearly amplify intensity of the spontaneous Raman noise of SRS, shown as Eqn (2) in which the intensity of Stokes line I s (ν s , L) is also decided by gain length L besides Raman noise I sn (ν s ), fluorescent seeds I seed (ν s ), gain coefficient g s , intensity of pumping light I p , and attenuation coefficient α. Moreover, comparing with the effect of I seed (ν s ), the gain length L in e-index decides distinctly on the intensity of Stokes line.…”

“…Therefore, in recent years, there are extensive applications in nonlinear optics, biological, and other fields. [1][2][3] However, the SRS spectrum of molecules is always a multistage cascade spectrum dominated by active mode with maximum Raman cross section. [4] The depletion of the pumping laser intensity by this SRS buildup prevents the significant amplification of SRS of other Raman active modes.…”