TY - JOUR
T1 - Direct Ultraviolet Laser-Induced Reduction of Disulfide Bonds in Insulin and Vasopressin
AU - Gammelgaard, Simon K
AU - Petersen, Steffen B
AU - Haselmann, Kim F
AU - Nielsen, Peter Kresten
N1 - Copyright © 2020 American Chemical Society.
PY - 2020/4/14
Y1 - 2020/4/14
N2 - Ultraviolet (UV) light has been shown to induce reduction of disulfide bonds in proteins in solution. The photoreduction is proposed to be a result of electron donation from excited Tyr or Trp residues. In this work, a powerful UV femtosecond laser was used to generate photoreduced products, while the hypothesis of Tyr/Trp mediation was studied with spectroscopy and mass spectrometry. With limited irradiation times of 3 min or less at 280 nm, the laser-induced reduction in arginine vasopressin and human insulin led to significant yields of ∼3% stable reduced product. The photogenerated thiols required acidic pH for stabilization, while neutral pH primarily caused scrambling and trisulfide formation. Interestingly, there was no direct evidence that Tyr/Trp mediation was a required criterion for the photoreduction of disulfide bonds. Intermolecular electron transfer remained a possibility for insulin but was ruled out for vasopressin. We propose that an additional mechanism should be increasingly considered in UV light-induced reduction of disulfide bonds in solution, in which a single UV photon is directly absorbed by the disulfide bond.
AB - Ultraviolet (UV) light has been shown to induce reduction of disulfide bonds in proteins in solution. The photoreduction is proposed to be a result of electron donation from excited Tyr or Trp residues. In this work, a powerful UV femtosecond laser was used to generate photoreduced products, while the hypothesis of Tyr/Trp mediation was studied with spectroscopy and mass spectrometry. With limited irradiation times of 3 min or less at 280 nm, the laser-induced reduction in arginine vasopressin and human insulin led to significant yields of ∼3% stable reduced product. The photogenerated thiols required acidic pH for stabilization, while neutral pH primarily caused scrambling and trisulfide formation. Interestingly, there was no direct evidence that Tyr/Trp mediation was a required criterion for the photoreduction of disulfide bonds. Intermolecular electron transfer remained a possibility for insulin but was ruled out for vasopressin. We propose that an additional mechanism should be increasingly considered in UV light-induced reduction of disulfide bonds in solution, in which a single UV photon is directly absorbed by the disulfide bond.
UR - http://www.scopus.com/inward/record.url?scp=85083357157&partnerID=8YFLogxK
U2 - 10.1021/acsomega.9b04375
DO - 10.1021/acsomega.9b04375
M3 - Journal article
C2 - 32309706
SN - 2470-1343
VL - 5
SP - 7962
EP - 7968
JO - ACS Omega
JF - ACS Omega
IS - 14
ER -