[Three dimensional structure of the dimeric gene-engineered variant of green fluorescent protein EGFP-K162Q in P6(1) crystal space group].

The crystal structure of the dimeric green fluorescent protein EGFP-K162Q with C-terminal deletion MDELYK (EGFPv) has been determined in space group P6 at resolution 1.34 A. The obtained structure has been compared with that of the monomeric form of EGFP (green biomarker with enhanced photophysical properties) determined in other crystal space group P2(1)2(1)2(1) at resolution 1.50 and 1.35 A [1, 2]. Two subunits in the EGFPv structure are packed at 75 degrees with the contact surface approximately 800 A2. The dimeric structure is stabilized by six hydrogen bonds and the central hydrophobic core built of six residues. The RMSD value for Calpha atoms of 3-230 residues in the superimposed P61 and P2(1)2(1)2(1) structures is 0.55 A. The distinguishing feature of EGFPv- P6(1) structure, compared with that of EGFP-P2(1)2(1)2(1), is the noticeable difference in orientation of the Glu222 side chain and also new conformation of the loop fragment 155-159 with deviations among the Calpha atoms of superimposed structures reaching for Lys156 - 4.6 A and Lys158 - 5.5 A

[Catalytic sites of medicinal leech enzyme destabilase-lysozyme (Mldl). Structure-functional correlation].

Based on three-dimensional model of the bifunctional enzyme Destabilase-Lysozyme (mlDL-Ds2) in complex with trimer of N-acetylglucosoamine (NAG)3 the functional role of the stereochemically based group of amino acids (Glu14, Asp26, Ser 29, Ser31, Lys38, His92), in manifestation of glycosidase and isopeptidase activities has been elucidated. By method of site-directed mutagenesis it has been shown that mlDL glycosidase active site includes catalytic Glu14 and Asp26, and isopeptidase site functions as Ser/Lys dyad presented by catalytic residues Lys38 and Ser29. Thus, among the invertebrate lysozymes mlDL presents first example of the bifunctional enzyme with identified position of the isopeptidase active site and localization of the corresponding catalytic residues.

[Three-dimensional structure of yellow fluorescent protein zYFP538 from Zoanthus sp. at the resolution 1.8 angstrom].

The three-dimensional structure of yellow fluorescent proteins zYFP538 (zFP538) from the button polyp Zoanthus sp. was determined at a resolution of 1.8 angstrom by X-ray analysis. The monomer of zYFP538 adopts a structure characteristic of the green fluorescent protein (GFP) family, a beta-barrel formed from 11 antiparallel beta segments and one internal alpha helix with a chromophore embedded into it. Like the TurboGFP, the beta-barrel of zYFP538 contains a water-filled pore leading to the chromophore Tyr67 residue, which presumably provides access of molecular oxygen necessary for the maturation process. The post-translational modification of the chromophore-forming triad Lys66-Tyr67-Gly68 results in a tricyclic structure consisting of a five-membered imidazolinone ring, a phenol ring of the Tyr67 residue, and an additional six-membered tetrahydropyridine ring. The chromophore formation is completed by cleavage of the protein backbone at the Calpha-N bond of Lys66. It was suggested that the energy conflict between the buried positive charge of the intact Lys66 side chain in the hydrophobic pocket formed by the Ile44, Leu46, Phe65, Leu204 and Leu219 side chains is the most probable trigger that induces the transformation of the bicyclic green form to the tricyclic yellow form. A stereochemical analysis of the contacting surfaces at the intratetramer interfaces helped reveal a group of conserved key residues responsible for the oligomerization. Along with others, these residues should be taken into account in designing monomeric forms suitable for practical application as markers of proteins and cell organelles.