title: Structural and kinetic characterization of early folding events in beta-lactoglobulin.
abstract: We have defined the structural and dynamic properties of an early folding intermediate of beta-lactoglobulin known to contain non-native alpha-helical structure. The folding of beta-lactoglobulin was monitored over the 100 micros--10 s time range using ultrarapid mixing techniques in conjunction with fluorescence detection and hydrogen exchange labeling probed by heteronuclear NMR. An initial increase in Trp fluorescence with a time constant of 140 micros is attributed to formation of a partially helical compact state. Within 2 ms of refolding, well protected amide protons indicative of stable hydrogen bonded structure were found only in a domain comprising beta-strands F, G and H, and the main alpha-helix, which was thus identified as the folding core of beta-lactoglobulin. At the same time, weak protection (up to approximately 10-fold) of amide protons in a segment spanning residues 12--21 is consistent with formation of marginally stable non-native alpha-helices near the N-terminus. Our results indicate that efficient folding, despite some local non-native structural preferences, is insured by the rapid formation of a native-like alpha/beta core domain.
authors.affiliation: Department of Physiology, School of Medicine, Gifu University, Tsukasa-machi, Gifu 500-8705, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Kuwata
authors.forename: R
authors.initials: R
authors.lastname: Shastry
authors.forename: H
authors.initials: H
authors.lastname: Cheng
authors.forename: M
authors.initials: M
authors.lastname: Hoshino
authors.forename: C A
authors.initials: CA
authors.lastname: Batt
authors.forename: Y
authors.initials: Y
authors.lastname: Goto
authors.forename: H
authors.initials: H
authors.lastname: Roder
mesh_terms: D000818:Animals; D002417:Cattle; D006859:Hydrogen / Q000378:metabolism; D006860:Hydrogen Bonding; D007700:Kinetics; D007782:Lactoglobulins / Q000737:chemistry* / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011489:Protein Denaturation; D017510:Protein Folding; D020673:Protein Renaturation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011522:Protons; D012997:Solvents; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics
chemical_list: D007782:Lactoglobulins; D011522:Protons; D012997:Solvents; D006859:Hydrogen