Intersubunit communication in tryptophan synthase by carbon-13 and fluorine-19 REDOR NMR

Biochemistry
L M McDowellJacob Schaefer

Abstract

The beta subunits of the 143-kDa alpha2beta2 tetrameric enzyme tryptophan synthase have been labeled by L-[ring-4-19F]phenylalanine and L-[phenol-4-13C]tyrosine in an effort to monitor the positions of these residues on ligand binding. Of the 13 phenylalanine and 11 tyrosine residues in the beta subunit, only three pairs have labels with 13C-19F separations of less than 6 angstrom. The beta subunit residues Tyr279 and Phe280 (each members of one of the three Tyr-Phe proximate pairs) have been suggested as possible conformational gates on ligand binding. The 188-MHz 19F NMR spectrum of the microcrystalline, double-labeled enzyme complex has five resolved lines under 5-kHz magic-angle spinning and 80-kHz proton dipolar decoupling. The distribution of beta-subunit 19F isotropic shifts is altered by addition of L-[3-13C]-serine to the mother liquor in contact with the microcrystals, consistent with a conformational rearrangement. The 13C label from serine is detected at 28 ppm as a methyl tautomer of bound aminoacrylate. The change in aromatic 19F chemical shifts on binding of serine indicates an alteration in local electric field gradients within the beta subunit. However, rotational-echo double-resonance 13C NMR (with 19F dephasi...Continue Reading

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Citations

Aug 10, 2007·Journal of Biomolecular NMR·W Trent FranksChad M Rienstra
Oct 1, 1996·Current Opinion in Structural Biology·L M McDowell, J Schaefer
Aug 12, 2004·Chemical Reviews·Stanley J Opella, Francesca M Marassi
Aug 1, 2002·Proceedings of the National Academy of Sciences of the United States of America·Chad M RienstraRobert G Griffin
Nov 11, 2015·Proceedings of the National Academy of Sciences of the United States of America·Andrew R BullerFrances H Arnold
Dec 25, 2007·Journal of Magnetic Resonance·Ulrich H N DürrAnne S Ulrich
May 4, 2007·Journal of Magnetic Resonance·Xingang ZhaoBrian D Sykes
Dec 24, 2002·Biophysical Journal·Stephan L GrageAnne S Ulrich
Sep 5, 2001·Journal of Magnetic Resonance·M L GilchristA E McDermott
Aug 1, 1997·Journal of Magnetic Resonance·J M Goetz, J Schaefer
Jun 19, 2004·Molecular Membrane Biology·Nielschr NielsenThomas Vosegaard
Oct 26, 2021·Chemical Reviews·Alexander A ShcherbakovMei Hong

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