Structure, dynamics, and insertion of a chloroplast targeting peptide in mixed micelles.
Nuclear-encoded, chloroplast-destined proteins are synthesized with transit sequences that contain all information to get them inside the organelle. Different proteins are imported via a general protein import machinery, but their transit sequences do not share amino acid homology. It has been suggested that interactions between transit sequence and chloroplast envelope membrane lipids give rise to recognizable, structural motifs. In this study a detailed investigation of the structural, dynamical, and topological features of an isolated transit peptide associated with mixed micelles is described. The structure of the preferredoxin transit peptide in these micelles was studied by circular dichroism (CD) and multidimensional NMR techniques. CD experiments indicated that the peptide, which is unstructured in aqueous solution, obtained helical structure in the presence of the micelles. By NMR it is shown that the micelles introduced ill-defined helical structures in the transit peptide. Heteronuclear relaxation experiments showed that the whole peptide backbone is very flexible. The least dynamic segments are two N- and C-terminal helical regions flanking an unstructured proline-rich amino acid stretch. Finally, the insertion of the peptide backbone in the hydrophobic interior of the micelle was investigated by use of hydrophobic spin-labels. The combined data result in a model of the transit peptide structure, backbone dynamics, and insertion upon its interaction with mixed micelles.
Wienk
H L
HL
Department of Biochemistry of Membranes, Centre of Biomembranes and Lipid Enzymology, Institute of Biomembranes, Utrecht, The Netherlands. h.l.j.wienk@chem.uu.nl
Wechselberger
R W
RW
Czisch
M
M
de Kruijff
B
B
D000595:Amino Acid Sequence; D002736:Chloroplasts / Q000737:chemistry*; D002942:Circular Dichroism; D005288:Ferredoxins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D019684:Magnoliopsida / Q000737:chemistry; D008823:Micelles; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D011487:Protein Conformation
D005288:Ferredoxins; D008823:Micelles; D010455:Peptides