pKa Determination of a Histidine Residue in a Short Peptide Using Raman Spectroscopy

Molecules

Anders Malmendal¹  Casey H. Londergan²  Brett H. Pogostin²  Karin S. Åkerfeldt² 

[1] Biochemistry and Structural Biology, Department of Chemistry, Lund University, PO Box 124, SE-221 00 Lund, Sweden;Department of Chemistry, Haverford College, Haverford, PA 19041, USA;

关键词: Raman spectroscopy;    1H NMR spectroscopy;    vibrational probes;    histidine;    peptides;    proteins;    isotopic labeling;    deuterium replacement;    acid dissociation constant;   

DOI  :  10.3390/molecules24030405

来源: DOAJ

【 摘 要 】

Determining the pKa of key functional groups is critical to understanding the pH-dependent behavior of biological proteins and peptide-based biomaterials. Traditionally, ¹H NMR spectroscopy has been used to determine the pKa of amino acids; however, for larger molecules and aggregating systems, this method can be practically impossible. Previous studies concluded that the C-D stretches in Raman are a useful alternative for determining the pKa of histidine residues. In this study, we report on the Raman application of the C2-D probe on histidine’s imidazole side chain to determining the pKa of histidine in a short peptide sequence. The pKa of the tripeptide was found via difference Raman spectroscopy to be 6.82, and this value was independently confirmed via ¹H NMR spectroscopy on the same peptide. The C2-D probe was also compared to other Raman reporters of the protonation state of histidine and was determined to be more sensitive and reliable than other protonation-dependent signals. The C2-D Raman probe expands the tool box available to chemists interested in directly interrogating the pKa’s of histidine-containing peptide and protein systems.

【 授权许可】

Unknown