3LVD

Crystal structure of GFP-like protein aceGFP_G222E (A. coerulescens). UV-photoconverted green form.

Summary for 3LVD

• Entry DOI: 10.2210/pdb3lvd/pdb
• Related: 3LVA 3LVC
• Descriptor: Green fluorescent protein, GLYCEROL (3 entities in total)
• Functional Keywords: green fluorescent protein, colorless gfp-like protein, chromophore biosynthesis, fluorescent protein
• Biological source: Aequorea coerulescens
• Total number of polymer chains: 2
• Total formula weight: 54088.70

Authors

Pletneva, N.V.,Pletnev, V.Z.,Pletnev, S.V. (deposition date: 2010-02-19, release date: 2010-03-09, Last modification date: 2024-10-09)

Primary citation

Pletneva, N.V.,Pletnev, V.Z.,Lukyanov, K.A.,Gurskaya, N.G.,Goryacheva, E.A.,Martynov, V.I.,Wlodawer, A.,Dauter, Z.,Pletnev, S.
Structural evidence for a dehydrated intermediate in green fluorescent protein chromophore biosynthesis.
J.Biol.Chem., 285:15978-15984, 2010

PubMed Abstract: The acGFPL is the first-identified member of a novel, colorless and non-fluorescent group of green fluorescent protein (GFP)-like proteins. Its mutant aceGFP, with Gly replacing the invariant catalytic Glu-222, demonstrates a relatively fast maturation rate and bright green fluorescence (lambda(ex) = 480 nm, lambda(em) = 505 nm). The reverse G222E single mutation in aceGFP results in the immature, colorless variant aceGFP-G222E, which undergoes irreversible photoconversion to a green fluorescent state under UV light exposure. Here we present a high resolution crystallographic study of aceGFP and aceGFP-G222E in the immature and UV-photoconverted states. A unique and striking feature of the colorless aceGFP-G222E structure is the chromophore in the trapped intermediate state, where cyclization of the protein backbone has occurred, but Tyr-66 still stays in the native, non-oxidized form, with C(alpha) and C(beta) atoms in the sp(3) hybridization. This experimentally observed immature aceGFP-G222E structure, characterized by the non-coplanar arrangement of the imidazolone and phenolic rings, has been attributed to one of the intermediate states in the GFP chromophore biosynthesis. The UV irradiation (lambda = 250-300 nm) of aceGFP-G222E drives the chromophore maturation further to a green fluorescent state, characterized by the conventional coplanar bicyclic structure with the oxidized double Tyr-66 C(alpha)=C(beta) bond and the conjugated system of pi-electrons. Structure-based site-directed mutagenesis has revealed a critical role of the proximal Tyr-220 in the observed effects. In particular, an alternative reaction pathway via Tyr-220 rather than conventional wild type Glu-222 has been proposed for aceGFP maturation.

PubMed: 20220148
DOI: 10.1074/jbc.M109.092320

Experimental method

X-RAY DIFFRACTION (1.751 Å)