Deuterium-Enhanced Raman Spectroscopy for Histidine pKa Determination in a pH-Responsive Hydrogel.
Biophys J
; 119(9): 1701-1705, 2020 11 03.
Article
em En
| MEDLINE
| ID: mdl-33080220
ABSTRACT
We report here a method for the determination of the pKa of histidine in complex or heterogeneous systems amenable to neither solid-state nor solution NMR spectroscopy. Careful synthesis of a fluorenylmethyloxycarbonyl- and trityl-protected, C2-deuterated histidine produces a vibrational-probe-equipped amino acid that can readily be incorporated into any peptide accessible by standard solid-phase methods. The frequency of the unique, Raman-active stretching vibration of this C2-D probe is a clear reporter of the protonation state of histidine. We investigate here a pH-sensitive peptide that self-assembles to form a hydrogel at neutral pH. The pKa of the lone histidine residue in the peptide, which is likely responsible for this pH-dependent behavior, cannot be investigated by NMR spectroscopy because of the supramolecular, soft nature of the gel. However, after synthesizing a C2-deuterated-histidine-containing peptide, we were able to follow the protonation state of histidine throughout a pH titration using Raman difference spectroscopy, thereby precisely determining the pKa of interest.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Análise Espectral Raman
/
Histidina
Idioma:
En
Revista:
Biophys J
Ano de publicação:
2020
Tipo de documento:
Article