Proline scanning mutagenesis reveals non-native fold in the molten globule state of equine beta-lactoglobulin.
The secondary structure in the molten globule state (an equilibrium analogue of a burst-phase folding intermediate) of equine beta-lactoglobulin was investigated by changes in the circular dichroic spectrum induced by a series of site-directed proline substitutions. The results challenge the structural picture obtained from previous hydrogen/deuterium exchange experiments. A stable non-native alpha-helix was found to exist in the region corresponding to the eighth strand (H strand) in the native structure, where the backbone amide protons are the most strongly protected from exchange. Therefore, the backbone topology in the folding core is significantly different from that in the native structure. This indicates that the burst-phase folding intermediate of beta-lactoglobulin is a trapped species because of misfolded backbone topology.
Nakagawa
Kanako
K
Department of Bioinformatics, Soka University, 1-236 Tangi-cho, Hachioji, Tokyo 192-8577, Japan.
Tokushima
Akihito
A
Fujiwara
Kazuo
K
Ikeguchi
Masamichi
M
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D000818:Animals; D002942:Circular Dichroism; D006736:Horses; D007782:Lactoglobulins / Q000737:chemistry* / Q000235:genetics*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D011392:Proline / Q000235:genetics*; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary / Q000235:genetics; D013816:Thermodynamics
D007782:Lactoglobulins; D011392:Proline