skip to main content
US FlagAn official website of the United States government
dot gov icon
Official websites use .gov
A .gov website belongs to an official government organization in the United States.
https lock icon
Secure .gov websites use HTTPS
A lock ( lock ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.


Title: Backbone 1H, 15N and 13C resonance assignments of the 27kDa fluorescent protein mCherry
Abstract mCherry is one of the most successfully applied monomeric red fluorescent proteins (RFPs) for in vivo and in vitro imaging. However, questions pertaining to the photostability of the RFPs remain and rational further engineering of their photostability requires information about the fluorescence quenching mechanism in solution. To this end, NMR spectroscopic investigations might be helpful, and we present the near-complete backbone NMR chemical shift assignment to aid in this pursuit.  more » « less
Award ID(s):
1734006
PAR ID:
10504208
Author(s) / Creator(s):
; ; ;
Publisher / Repository:
Biomolecular NMR Assignments
Date Published:
Journal Name:
Biomolecular NMR Assignments
Volume:
17
Issue:
2
ISSN:
1874-2718
Page Range / eLocation ID:
243 to 247
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract Red fluorescent proteins (RFPs) represent an increasingly popular class of genetically encodable bioprobes and biomarkers that can advance next‐generation breakthroughs across the imaging and life sciences. Since the rational design of RFPs with improved functions or enhanced versatility requires a mechanistic understanding of their working mechanisms, while fluorescence is intrinsically an ultrafast event, a suitable toolset involving steady‐state and time‐resolved spectroscopic techniques has become powerful in delineating key structural features and dynamic steps which govern irreversible photoconverting or reversible photoswitching RFPs, and large Stokes shift (LSS)RFPs. The pertinentcis‐transisomerization and protonation state change of RFP chromophores in their local environments, involving key residues in protein matrices, lead to rich and complicated spectral features across multiple timescales. In particular, ultrafast excited‐state proton transfer in various LSSRFPs showcases the resolving power of wavelength‐tunable femtosecond stimulated Raman spectroscopy (FSRS) in mapping a photocycle with crucial knowledge about the red‐emitting species. Moreover, recent progress in noncanonical RFPs with a site‐specifically modified chromophore provides an appealing route for efficient engineering of redder and brighter RFPs, highly desirable for bioimaging. Such an effective feedback loop involving physical chemists, protein engineers, and biomedical microscopists will enable future successes to expand fundamental knowledge and improve human health. 
    more » « less
  2. We study single- and multiple-ratio robust fractional 0-1 programming problems (RFPs). In particular, this work considers RFPs under a wide range of disjoint and joint uncertainty sets, where the former implies separate uncertainty sets for each numerator and denominator and the latter accounts for different forms of interrelatedness between them. We first demonstrate that unlike the deterministic case, a single-ratio RFP is nondeterministic polynomial-time hard under general polyhedral uncertainty sets. However, if the uncertainty sets are imbued with a certain structure, variants of the well-known budgeted uncertainty, the disjoint and joint single-ratio RFPs are polynomially solvable when the deterministic counterpart is. We also propose mixed-integer linear programming (MILP) formulations for multiple-ratio RFPs. We conduct extensive computational experiments using test instances based on real and synthetic data sets to evaluate the performance of our MILP reformulations as well as to compare the disjoint and joint uncertainty sets. Finally, we demonstrate the value of the robust approach by examining the robust solution in a deterministic setting and vice versa. 
    more » « less
  3. Abstract Recent advances in sustainable optoelectronics including photovoltaics, light‐emitting diodes, transistors, and semiconductors have been enabled by π‐conjugated organic molecules. A fundamental understanding of light‐matter interactions involving these materials can be realized by time‐resolved electronic and vibrational spectroscopies. In this Minireview, the photoinduced mechanisms including charge/energy transfer, electronic (de)localization, and excited‐state proton transfer are correlated with functional properties encompassing optical absorption, fluorescence quantum yield, conductivity, and photostability. Four naturally derived molecules (xylindein, dimethylxylindein, alizarin, indigo) with ultrafast spectral insights showcase efficient energy dissipation involving H‐bonding networks and proton motions, which yield high photostability. Rational design principles derived from such investigations could increase the efficiency for light harvesting, triplet formation, and photosensitivity for improved and versatile optoelectronic performance. 
    more » « less
  4. A new deep-blue emitting and highly fluorescent anthracene (ANTH) derivative containing perfluorobenzyl (BnF) groups, 9,10-ANTH(BnF)2, was synthesized in a single step reaction of ANTH or ANTH(Br)2with BnFI, using either a high-temperature Cu-/Na2S2O3-promoted reaction or via a room-temperature photochemical reaction. Its structure was elucidated by NMR spectroscopy and single crystal X-ray diffractometry. The latter revealed no π–π interaction between neighboring ANTH cores. A combination of high photoluminescence quantum yield (PLQY) of 0.85 for 9,10-ANTH(BnF)2, its significantly improved photostability compared to ANTH and 9,10-ANTH derivatives, and a simple synthetic access makes it an attractive material as a deep-blue OLED emitter and an efficient fluorescent probe. 
    more » « less
  5. Abstract Large Stokes shift (LSS) red fluorescent proteins (RFPs) are highly desirable for bioimaging advances. The RFP mKeima, with coexistingcis‐andtrans‐isomers, holds significance as an archetypal system for LSS emission due to excited‐state proton transfer (ESPT), yet the mechanisms remain elusive. We implemented femtosecond stimulated Raman spectroscopy (FSRS) and various time‐resolved electronic spectroscopies, aided by quantum calculations, to dissect thecis‐ andtrans‐mKeima photocycle from ESPT, isomerization, to ground‐state proton transfer in solution. This work manifests the power of FSRS with global analysis to resolve Raman fingerprints of intermediate states. Importantly, the deprotonatedtrans‐isomer governs LSS emission at 620 nm, while the deprotonatedcis‐isomer's 520 nm emission is weak due to an ultrafastcis‐to‐transisomerization. Complementary spectroscopic techniques as a table‐top toolset are thus essential to study photochemistry in physiological environments. 
    more » « less