Solution structure of a lipid transfer protein extracted from rice seeds. Comparison with homologous proteins.

Nuclear magnetic resonance (NMR) spectroscopy was used to determine the three dimensional structure of rice nonspecific lipid transfer protein (ns-LTP), a 91 amino acid residue protein belonging to the broad family of plant ns-LTP. Sequence specific assignment was obtained for all but three HN backbone H-1 resonances and for more than 95% of the H-1 side-chain resonances using a combination of H-1 2D NOESY; TOCSY and COSY experiments at 293 K. The structure was calculated on the basis of four disulfide bridge restraints, 1259 distance constraints derived from H-1-H-1 Overhauser effects, 72 phi angle restraints and 32 hydrogen-bond restraints. The final solution structure involves four helices (H1.: Cys3-Arg18, H2: Ala25-Ala37, H3: Thr41-Ala54 and H4: Ala66-Cys73) followed by a long C-terminal tail (T) with no observable regular structure. N-capping residues (Thr2, Ser24, Thr-40), whose sidechain oxygen atoms are involved in hydrogen bonds with i + 3 amide proton additionally stabilize the N termini of the first three helices. The fourth helix involving Pro residues display a mixture of alpha and 3(10) conformation. The rms deviation of 14 final structures with respect to the average structure is 1.14 +/- 0.16 Angstrom for all heavy atoms (C, N, O and S) and 0.72 +/- 0.01 Angstrom for the backbone atoms. The global fold of rice ns-LTP is close to the previously published structures of wheat, barley and maize ns-LTPs exhibiting nearly identical pattern of the numerous sequence specific interactions. As reported previously for different four-helix topology proteins, hydrophobic, hydrogen bonding and electrostatic mechanisms of fold stabilization were found for the lice ns-LTP. The sequential alignment of 36 ns-LTP primary structures strongly suggests that there is a uniform pattern of specific long-range interactions tin terms of sequence), which stabilize the fold of all plant ns-LTPs.