TY - JOUR
T1 - Structural Characterization of Acylimine-Containing Blue and Red Chromophores in mTagBFP and TagRFP Fluorescent Proteins
AU - Subach, Oksana M.
AU - Malashkevich, Vladimir N.
AU - Zencheck, Wendy D.
AU - Morozova, Kateryna S.
AU - Piatkevich, Kiryl D.
AU - Almo, Steven C.
AU - Verkhusha, Vladislav V.
N1 - Funding Information:
This work was supported by the National Institutes of Health (grant GM073913 to V.V.V.) and by the Albert Einstein Cancer Center. We thank H.Xiao for measuring mass spectra and U. Ramagopal for assistance with structure determination. We also thank the staff of the X29 beamline at the National Synchrotron Light Source, Brookhaven National Laboratory.
PY - 2010/4/23
Y1 - 2010/4/23
N2 - We determined the 2.2 Å crystal structures of the red fluorescent protein TagRFP and its derivative, the blue fluorescent protein mTagBFP. The crystallographic analysis is consistent with a model in which TagRFP has the trans coplanar anionic chromophore with the conjugated π-electron system, similar to that of DsRed-like chromophores. Refined conformation of mTagBFP suggests the presence of an N-acylimine functionality in its chromophore and single Cα-Cβ bond in the Tyr64 side chain. Mass spectrum of mTagBFP chromophore-bearing peptide indicates a loss of 20 Da upon maturation, whereas tandem mass spectrometry reveals that the Cα-N bond in Leu63 is oxidized. These data indicate that mTagBFP has a new type of the chromophore, N-[(5-hydroxy-1H-imidazole-2-yl)methylidene]acetamide. We propose a chemical mechanism in which the DsRed-like chromophore is formed via the mTagBFP-like blue intermediate.
AB - We determined the 2.2 Å crystal structures of the red fluorescent protein TagRFP and its derivative, the blue fluorescent protein mTagBFP. The crystallographic analysis is consistent with a model in which TagRFP has the trans coplanar anionic chromophore with the conjugated π-electron system, similar to that of DsRed-like chromophores. Refined conformation of mTagBFP suggests the presence of an N-acylimine functionality in its chromophore and single Cα-Cβ bond in the Tyr64 side chain. Mass spectrum of mTagBFP chromophore-bearing peptide indicates a loss of 20 Da upon maturation, whereas tandem mass spectrometry reveals that the Cα-N bond in Leu63 is oxidized. These data indicate that mTagBFP has a new type of the chromophore, N-[(5-hydroxy-1H-imidazole-2-yl)methylidene]acetamide. We propose a chemical mechanism in which the DsRed-like chromophore is formed via the mTagBFP-like blue intermediate.
KW - CHEMBIO
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U2 - 10.1016/j.chembiol.2010.03.005
DO - 10.1016/j.chembiol.2010.03.005
M3 - Article
C2 - 20416505
AN - SCOPUS:77950951096
SN - 1074-5521
VL - 17
SP - 333
EP - 341
JO - Chemistry and Biology
JF - Chemistry and Biology
IS - 4
ER -