학술논문
Stalling chromophore synthesis of the fluorescent protein Venus reveals the molecular basis of the final oxidation step.
Document Type
Academic Journal
Author
Auhim HS; School of Biosciences, Molecular Biosciences Division, Cardiff University Sir Martin Evans Building Cardiff CF10 3AX UK jonesdd@cardiff.ac.uk +44 (0)29 2087 4290.; Department of Biology, College of Science, University of Baghdad Baghdad Iraq.; Grigorenko BL; Chemistry Department, Lomonosov Moscow State University Leninskie Gory, 1-3 Moscow Russian Federation anem@lcc.chem.msu.ru +7 495 939 1096.; Emanuel Institute of Biochemical Physics, Russian Academy of Sciences Moscow Russian Federation.; Harris TK; School of Biosciences, Molecular Biosciences Division, Cardiff University Sir Martin Evans Building Cardiff CF10 3AX UK jonesdd@cardiff.ac.uk +44 (0)29 2087 4290.; Aksakal OE; School of Biosciences, Molecular Biosciences Division, Cardiff University Sir Martin Evans Building Cardiff CF10 3AX UK jonesdd@cardiff.ac.uk +44 (0)29 2087 4290.; Polyakov IV; Chemistry Department, Lomonosov Moscow State University Leninskie Gory, 1-3 Moscow Russian Federation anem@lcc.chem.msu.ru +7 495 939 1096.; Emanuel Institute of Biochemical Physics, Russian Academy of Sciences Moscow Russian Federation.; Berry C; School of Biosciences, Molecular Biosciences Division, Cardiff University Sir Martin Evans Building Cardiff CF10 3AX UK jonesdd@cardiff.ac.uk +44 (0)29 2087 4290.; Gomes GDP; Department of Chemistry, University of Toronto 80 St. George Street Toronto ON M5S 3H6 Canada.; Department of Computer Science, University of Toronto 214 College St. Toronto Ontario M5T 3A1 Canada.; Alabugin IV; Department of Chemistry and Biochemistry, Florida State University Tallahassee Fl 32306 USA alabugin@chem.fsu.edu +1 850 644 5795.; Rizkallah PJ; School of Medicine, Cardiff University CF14 4XN UK.; Nemukhin AV; Chemistry Department, Lomonosov Moscow State University Leninskie Gory, 1-3 Moscow Russian Federation anem@lcc.chem.msu.ru +7 495 939 1096.; Emanuel Institute of Biochemical Physics, Russian Academy of Sciences Moscow Russian Federation.; Jones DD; School of Biosciences, Molecular Biosciences Division, Cardiff University Sir Martin Evans Building Cardiff CF10 3AX UK jonesdd@cardiff.ac.uk +44 (0)29 2087 4290.
Source
Publisher: Royal Society of Chemistry Country of Publication: England NLM ID: 101545951 Publication Model: Electronic Cited Medium: Print ISSN: 2041-6520 (Print) Linking ISSN: 20416520 NLM ISO Abbreviation: Chem Sci Subsets: PubMed not MEDLINE
Subject
Language
English
ISSN
2041-6520
Abstract
Fluorescent proteins (FPs) have revolutionised the life sciences, but the mechanism of chromophore maturation is still not fully understood. Here we show that incorporation of a photo-responsive non-canonical amino acid within the chromophore stalls maturation of Venus, a yellow FP, at an intermediate stage; a crystal structure indicates the presence of O 2 located above a dehydrated enolate form of the imidazolone ring, close to the strictly conserved Gly67 that occupies a twisted conformation. His148 adopts an "open" conformation so forming a channel that allows O 2 access to the immature chromophore. Absorbance spectroscopy supported by QM/MM simulations suggests that the first oxidation step involves formation of a hydroperoxyl intermediate in conjunction with dehydrogenation of the methylene bridge. A fully conjugated mature chromophore is formed through release of H 2 O 2 , both in vitro and in vivo . The possibility of interrupting and photochemically restarting chromophore maturation and the mechanistic insights open up new approaches for engineering optically controlled fluorescent proteins.
Competing Interests: There are no conflicts to declare.
(This journal is © The Royal Society of Chemistry.)
Competing Interests: There are no conflicts to declare.
(This journal is © The Royal Society of Chemistry.)