1h6c: Difference between revisions

The NADP(H)-dependent enzyme glucose-fructose oxidoreductase (GFOR) is a, classic example of a redox protein that is translocated across a membrane, in fully folded form. GFOR is synthesized in the cytoplasm with a, 52-residue signal peptide, giving a precursor form, preGFOR, that is fully, active and has its cofactor tightly bound. A twin-arginine motif in the, signal peptide directs it to a Sec-independent pathway by which it is, translocated, in fully folded form, into the periplasm where it functions, to produce sorbitol for osmoprotection. We have determined the crystal, structures of four different forms of preGFOR, (i) oxidized preGFOR, with, succinate bound in the active site, (ii) oxidized preGFOR with glycerol, bound, (iii) reduced preGFOR in 0.3 M glucose, and (iv) reduced preGFOR in, 1.5 M sorbitol, at resolutions of 2.2, 2.05, 2.5, and 2.6 A, respectively., In all four crystal structures, the signal peptide is disordered, implying, a flexibility that may be important for its interaction with the, translocation apparatus; a factor contributing to this disorder may be the, high positive charge of the protein surface in the region where the signal, peptide emerges. This may disfavor a stable association between the signal, peptide and the rest of the protein. The crystal structures show that the, mature enzyme portion of preGFOR is identical to native GFOR, in structure, and cofactor binding, explaining the enzymatic activity of the precursor, form. In the glycerol complex, preGFOR(gll), a bound glycerol molecule, models the binding of the glucose substrate, with its O1 atom hydrogen, bonded to the essential acid/base catalyst, Tyr269, and C1 only 3 A from, C4 of the nicotinamide. In the glucose-soaked structure, preGFOR(glu), we, identify a conformational change of the nearby Lys181 that probably, results from the oxidation of glucose to gluconolactone, and functions to, prevent rebinding of glucose prior to the binding of fructose. In this, conformational change, the Lys181 side chain moves closer to the, nicotinamide ring, stabilized by its increased negative charge.

1H6C is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Zymomonas_mobilis Zymomonas mobilis] with NDP, SIN and GOL as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Glucose--fructose_oxidoreductase Glucose--fructose oxidoreductase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.1.99.28 1.1.99.28] Structure known Active Site: AC1. Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1H6C OCA].  

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