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.
journal_name
Biophys Jjournal_title
Biophysical journalauthors
Braun GA,Pogostin BH,Pucetaite M,Londergan CH,Åkerfeldt KSdoi
10.1016/j.bpj.2020.09.011subject
Has Abstractpub_date
2020-11-03 00:00:00pages
1701-1705issue
9eissn
0006-3495issn
1542-0086pii
S0006-3495(20)30721-9journal_volume
119pub_type
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