title: Core mutants of the immunoglobulin binding domain of streptococcal protein G: stability and structural integrity.
abstract: A library of core mutants of the GB1 domain of streptococcal protein G was created, and the structure and stability of selected members was assessed by 1H-15N heteronuclear correlation NMR spectroscopy and fluorescence. All mutants comprised changes in beta-sheet residues, with sidechains at positions 5 (Leu), 7 (Leu), 52 (Phe) and 54 (Val) forming the beta-sheet side of the sheet-helix core interface. A solvent exposed position Ile-6 was chosen as a control. Randomization of bases at codon positions 1 and 3 with thymine at position 2 introduces five possible hydrophobic amino acids, namely Leu, Val, Ile, Phe, and Met. The distribution of encoded amino acids at all five positions is approximately as expected theoretically and indicates that no major bias was introduced towards particular residues. The overall structural integrity of several mutants, as assessed by NMR, ranges from very close to wild type to fully unfolded. Interestingly, the stability of the mutants is not strictly correlated with the number of changes or residue volume.
authors.affiliation: Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520, USA. gronenborn@vger.niddk.nih.gov
authors.forename: A M
authors.initials: AM
authors.lastname: Gronenborn
authors.forename: M K
authors.initials: MK
authors.lastname: Frank
authors.forename: G M
authors.initials: GM
authors.lastname: Clore
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites; D007074:Immunoglobulin G / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013816:Thermodynamics
chemical_list: D001426:Bacterial Proteins; C034284:IgG Fc-binding protein, Streptococcus; D007074:Immunoglobulin G