Biophysical Journal 64: 492-501 (1993)
© 1993 the Biophysical Society

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Solid state ³¹P cross-polarization/magic angle sample spinning nuclear magnetic resonance studies of crystalline glycogen phosphorylase b

Jocelyn E. Taguchi ^*, Stephen J. Heyes ^*, David Barford , Louise N. Johnson  and Christopher M. Dobson ^*

Inorganic Chemistry Laboratory, Oxford Centre for Molecular Sciences, University of Oxford, Oxford, United Kingdom
Laboratory of Molecular Biophysics, Oxford Centre for Molecular Sciences, University of Oxford, Oxford, United Kingdom

ABSTRACT

³¹P cross-polarization/magic angle sample spinning nuclear magnetic resonance spectra have been obtained for pyridoxal 5'-phosphate (PLP) bound to glycogen phosphorylase b (GPb) in two different crystalline forms, monoclinic and tetragonal. Analysis of the intensities of the spinning sidebands in the nuclear magnetic resonance spectra has enabled estimates of the principal values of the ³¹P chemical shift tensors to be obtained. Differences between the two sets of values suggest differences in the environment of the phosphate moiety of the pyridoxal phosphate in the two crystalline forms. The tensor for the tetragonal crystalline form, T state GPb, is fully consistent with those found for dianionic phosphate groups in model compounds. The spectrum for the monoclinic crystalline form, R state GPb, although closer to that of dianionic than monoanionic model phosphate compounds, deviates significantly from that expected for a simple dianion or monoanion. This is likely to result from specific interactions between the PLP phosphate group and residues in its binding site in the protein. A possible explanation for the spectrum of the monoclinic crystals is that the shift tensor is averaged by a proton exchange process between different ionization states of the PLP associated with the presence of a sulfate ion bound in the vicinity of the PLP.