title: Folding and functional complementation of engineered fragments from yeast phosphoglycerate kinase.
abstract: A set of protein fragments was produced by site-directed mutagenesis followed by chemical cleavage of phosphoglycerate kinase according to a previously described method [Pecorari et al. (1993) Protein Eng. 6, 313-325]. The cleavage positions were chosen in order to correspond to limits between structural subdomains. These isolated fragments were studied by circular dichroism, folding transitions, and cross-linking analyses. It appears that fragments corresponding to globular subdomains in the protein can recover the expected helix content. However, the cooperativity classically observed in the folding transitions of natural proteins is only observed for fragments larger than a domain. Previous studies have shown that the isolated C-terminal domain is an autonomous folding unit which displays a single cooperatine transition [Missiakas et al. (1990) Biochemistry 29, 8683-8689]. The results presented here show that the presence in a fragment of a sequence overpassing that of the C-terminal domain modifies its folding process. Reassociation experiments suggest that the efficiency of the complementation process is not related to the folding autonomy of the isolated fragments.
authors.affiliation: Laboratoire d'Enzymologie physico-chimique et moléculaire, Unité associée du Centre National de la Recherche Scientifique, Universite de Paris-sud, Orsay, France.
authors.forename: F
authors.initials: F
authors.lastname: Pecorari
authors.forename: C
authors.initials: C
authors.lastname: Guilbert
authors.forename: P
authors.initials: P
authors.lastname: Minard
authors.forename: M
authors.initials: M
authors.lastname: Desmadril
authors.forename: J M
authors.initials: JM
authors.lastname: Yon
mesh_terms: D002942:Circular Dichroism; D003432:Cross-Linking Reagents / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry; D008970:Molecular Weight; D010735:Phosphoglycerate Kinase / Q000737:chemistry*; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D013696:Temperature; D013816:Thermodynamics; D015003:Yeasts / Q000201:enzymology
chemical_list: D003432:Cross-Linking Reagents; D005656:Fungal Proteins; D010735:Phosphoglycerate Kinase