title: Magic-angle spinning solid-state NMR spectroscopy of the beta1 immunoglobulin binding domain of protein G (GB1): 15N and 13C chemical shift assignments and conformational analysis.
abstract: Magic-angle spinning solid-state NMR (SSNMR) studies of the beta1 immunoglobulin binding domain of protein G (GB1) are presented. Chemical shift correlation spectra at 11.7 T (500 MHz 1H frequency) were employed to identify signals specific to each amino acid residue type and to establish backbone connectivities. High sensitivity and resolution facilitated the detection and assignment of every 15N and 13C site, including the N-terminal (M1) 15NH3, the C-terminal (E56) 13C', and side-chain resonances from residues exhibiting fast-limit conformational exchange near room temperature. The assigned spectra lend novel insight into the structure and dynamics of microcrystalline GB1. Secondary isotropic chemical shifts report on conformation, enabling a detailed comparison of the microcrystalline state with the conformation of single crystals and the protein in solution; the consistency of backbone conformation in these three preparations is the best among proteins studied so far. Signal intensities and line widths vary as a function of amino acid position and temperature. High-resolution spectra are observed near room temperature (280 K) and at <180 K, whereas resolution and sensitivity greatly degrade substantially near 210 K; the magnitude of this effect is greatest among the side chains of residues at the intermolecular interface of the microcrystal lattice, which we attribute to intermediate-rate translational diffusion of solvent molecules near the glass transition. These features of GB1 will enable its use as an excellent model protein not only for SSNMR methods development but also for fundamental studies of protein thermodynamics in the solid state.
authors.affiliation: Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, Illinois 61801, USA.
authors.forename: W Trent
authors.initials: WT
authors.lastname: Franks
authors.forename: Donghua H
authors.initials: DH
authors.lastname: Zhou
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Wylie
authors.forename: Brian G
authors.initials: BG
authors.lastname: Money
authors.forename: Daniel T
authors.initials: DT
authors.lastname: Graesser
authors.forename: Heather L
authors.initials: HL
authors.lastname: Frericks
authors.forename: Gurmukh
authors.initials: G
authors.lastname: Sahota
authors.forename: Chad M
authors.initials: CM
authors.lastname: Rienstra
mesh_terms: D000595:Amino Acid Sequence; D002247:Carbon Isotopes; D018360:Crystallography, X-Ray / Q000379:methods; D007136:Immunoglobulins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry*; D009587:Nitrogen Isotopes; D011485:Protein Binding; D011487:Protein Conformation; D012680:Sensitivity and Specificity; D013816:Thermodynamics; D013997:Time Factors
chemical_list: D002247:Carbon Isotopes; C029152:G-substrate; D007136:Immunoglobulins; D009419:Nerve Tissue Proteins; D009587:Nitrogen Isotopes