title: Structural and energetic consequences of disruptive mutations in a protein core.
abstract: We have characterized the properties of a set of variants of the N-terminal domain of lambda repressor bearing disruptive mutations in the hydrophobic core. These mutations include some that dramatically alter the total core residue volume (by up to six methylene groups) and some that place a single polar residue into the otherwise hydrophobic core. The structural properties of the purified proteins have been studied by CD spectroscopy, biological activity, recognition by conformation-specific monoclonal antibodies, and 1H NMR spectroscopy. The stabilities of the proteins have been measured by thermal and guanidine hydrochloride denaturation. Proteins with disruptive core mutations are found to display a continuum of increasingly nonnative properties. Large internal volume changes cause both significant conformational rearrangements and destabilization by up to 5 kcal/mol. Variants with polar substitutions at core positions no longer behave like well-folded proteins but rather display characteristics of molten globules. However, even proteins bearing some of the most disruptive mutations retain many of the crude secondary and tertiary structural features of the wild-type protein. These results indicate that primitive elements of native structure can form in the absence of normal core packing.
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
authors.forename: W A
authors.initials: WA
authors.lastname: Lim
authors.forename: D C
authors.initials: DC
authors.lastname: Farruggio
authors.forename: R T
authors.initials: RT
authors.lastname: Sauer
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D002942:Circular Dichroism; D004268:DNA-Binding Proteins; D019791:Guanidine; D006146:Guanidines / Q000494:pharmacology; D006358:Hot Temperature; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011487:Protein Conformation; D011489:Protein Denaturation; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000648:ultrastructure; D012995:Solubility; D013329:Structure-Activity Relationship; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins
chemical_list: D000911:Antibodies, Monoclonal; D004268:DNA-Binding Proteins; D006146:Guanidines; D012097:Repressor Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins; D019791:Guanidine