The molecular structure of green fluorescent protein

Yang, Fan; Moss, Larry G.; Phillips, George N.

doi:10.1038/nbt1096-1246

Cited by 1,446 publications

(1,289 citation statements)

References 39 publications

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“…This finding indicates that the recognition of the processed N-terminus, lacking the first three residues of the PEXEL (RxL), may be occurring at the PVM by a putative translocase and the inability of the parasite to traffic KAHRP(1-60)-GFP beyond the PVM could be due to a defect in this proposed recognition event. The crystal structure of GFP shows that the first nine amino acids are involved in the formation of an α-helix that plays an integral part in capping one end of the protein and may aid in folding events or in protecting the fluorophore [32,33]. It is possible that the close proximity of the processed PEXEL to this N-terminal α-helical cap of GFP obstructs the recognition of the processed N-terminus of the KAHRP chimera by a putative PVM translocase.…”

Section: Transport Across the Pvm Mediated By The Recognition Of The mentioning

confidence: 99%

N-terminal processing of proteins exported by malaria parasites

Chang

Falick

Carlton

et al. 2008

Molecular and Biochemical Parasitology

169

175

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Malaria parasites utilize a short N-terminal amino acid motif termed the Plasmodium export element (PEXEL) to export an array of proteins to the host erythrocyte during blood stage infection. Using immunoaffinity chromatography and mass spectrometry, insight into this signalmediated trafficking mechanism was gained by discovering that the PEXEL motif is cleaved and N-acetylated. PfHRPII and PfEMP2 are two soluble proteins exported by Plasmodium falciparum that were demonstrated to undergo PEXEL cleavage and N-acetylation, thus indicating that this Nterminal processing may be general to many exported soluble proteins. It was established that PEXEL processing occurs upstream of the brefeldin A-sensitive trafficking step in the P. falciparum secretory pathway, therefore cleavage and N-acetylation of the PEXEL motif occurs in the endoplasmic reticulum (ER) of the parasite. Furthermore, it was shown that the recognition of the processed N-terminus of exported proteins within the parasitophorous vacuole may be crucial for protein transport to the host erythrocyte. It appears that the PEXEL may be defined as a novel ER peptidase cleavage site and a classical N-acetyltransferase substrate sequence.

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Section: Transport Across the Pvm Mediated By The Recognition Of The mentioning

confidence: 99%

N-terminal processing of proteins exported by malaria parasites

Chang

Falick

Carlton

et al. 2008

Molecular and Biochemical Parasitology

169

175

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“…The green fluorescent protein, in Pacific jellyfish and similar animals, is another striking example of the use of amino acids to self-catalytically generate a useful protein-bound group. The fluorophore of the Escherichia coliproduced protein results from cyclization through condensation of the backbone carbonyl carbon of Ser 65 with the backbone amide nitrogen of Gly 67, and oxidation of the C,-Cp bond of Tyr 66 (Yang et al, 1996). A final interesting example of a redox-active, modified aminoacyl group is provided by NADH peroxidase of Enterococcus fueculis lOCl.…”

Section: Introductionmentioning

confidence: 99%

Covalent attachment of flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) to enzymes: The current state of affairs

1998

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The first identified covalent flavoprotein, a component of mammalian succinate dehydrogenase, was reported 42 years ago. Since that time, more than 20 covalent flavoenzymes have been described, each possessing one of five modes of FAD or FMN linkage to protein. Despite the early identification of covalent flavoproteins, the mechanisms of covalent bond formation and the roles of the covalent links are only recently being appreciated. The main focus of this review is, therefore, one of mechanism and function, in addition to surveying the types of linkage observed and the methods employed for their identification. Case studies are presented for a variety of covalent flavoenzymes, from which general findings are beginning to emerge.Keywords: covalent flavoproteins; flavin adenine dinucleotide; flavin mononucleotide; flavinylation; redox cofactors Cofactors are recruited by protein molecules to extend the chemistry available within the active sites of enzymes. The chemistries displayed are variable and the range of protein-specific cofactors available bears testimony to the types of biological reaction achieved by many enzyme molecules. Cofactors may be present in the freely dissociable form, but others, such as lipoic acid and biotin, are permanently linked to the host protein. Other cofactors (e.g., heme and pyridoxyl phosphate) fall into both categories in that they can be attached covalently to the protein or operate in the freely dissociable form. The covalent addition of specific cofactors to unique residues within a polypeptide chain is a fundamental problem in studies of biological molecular recognition. In many cases, this specificity is purchased at the expense of recruiting modification enzymes that direct the covalent addition of a cofactor to the host protein. For example, the additions of biotin and lipoic acid to the €-amino groups of specific Lys residues in carboxyltransferases and the 2-oxoacid dehydrogenases are directed by a biotin holoenzyme synthase (Sweetman et al., 1985;Takai et al., 1987) and Reprint requests to: N.S. Scmtton, Department of Biochemistry, University of Leicester, Adrian Building, University Road, Leicester LE1 7RH UK; e-mail: nss4@le.ac.uk; or W.S. McIntire, Molecular Biology Division, Department of Veterans Affairs Medical Center, San Francisco, California 94121; e-mail: wsm@itsa.ucsf.edu.lipoate-protein ligases (Brookfield et al., 1991; Moms et al., 1994), respectively. In a similar fashion, a heme cytochrome c lyase (Steiner et al., 1996) is responsible for the covalent addition of heme to two Cys residues via thioether linkages. In some enzymes, an existing side chain is modified to yield what is in effect a covalently attached cofactor, although no pre-existing cofactor molecule is incorporated into the enzyme. Several examples of this type of modification are provided in the next paragraph.For histidine decarboxylase of Lactobacillus 30A (Recsei & Snell, 1984), a pyruvoyl group is generated from an existing residue, Ser 82. The enzyme is synthesized as a proenzym...

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“…The off-state structure was cryogenically trapped after full conversion of crystals with green illumination 5 . The chromophore of Dronpa is derived from the Cys-Tyr-Gly tripeptide and is present in a cis conformation in the on state 3,4 , as in the A. victoria GFP 7,8 . Substantial structural rearrangements of the protein environment were shown to coincide with the cis-trans photoisomerization of the chromophore.…”

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confidence: 99%

Ground-state proton transfer in the photoswitching reactions of the fluorescent protein Dronpa

et al. 2013

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The reversible photoswitching between the 'on' and 'off' states of the fluorescent protein Dronpa involves photoisomerization as well as protein side-chain rearrangements, but the process of interconversion remains poorly characterized. Here we use time-resolved infrared measurements to monitor the sequence of these structural changes, but also of proton transfer events, which are crucial to the development of spectroscopic contrast. Light-induced deprotonation of the chromophore phenolic oxygen in the off state is a thermal ground-state process, which follows ultrafast (9 ps) trans-cis photoisomerization, and so does not involve excited-state proton transfer. Steady-state infrared difference measurements exclude protonation of the imidazolinone nitrogen in both the on and off states. Pump-probe infrared measurements of the on state reveal a weakening of the hydrogen bonding between Arg66 and the chromophore C ¼ O, which could be central to initiating structural rearrangement of Arg66 and His193 coinciding with the low quantum yield cis-trans photoisomerization

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The molecular structure of green fluorescent protein

Cited by 1,446 publications

References 39 publications

N-terminal processing of proteins exported by malaria parasites

N-terminal processing of proteins exported by malaria parasites

Covalent attachment of flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) to enzymes: The current state of affairs

Ground-state proton transfer in the photoswitching reactions of the fluorescent protein Dronpa

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