Determination of the three-dimensional solution structure of Raphanus sativus antifungal protein 1 by super(1)H NMR

Raphanus sativus Antifungal Protein 1 (Rs-AFP1) is a 51 amino acid residue plant defensin isolated from radish (Raphanus sativus L.) seeds. The three-dimensional structure in aqueous solution has been determined from two-dimensional super(1)H NMR data recorded at 500 MHz using the DIANA/REDAC calcul...

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Bibliographic Details
Published in:Journal of molecular biology 1998-01, Vol.279 (1), p.257-270
Main Authors: Fant, F, Vranken, W, Broekaert, W, Borremans, F
Format: Article
Language:English
Subjects:
Raphanus sativus
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Summary:Raphanus sativus Antifungal Protein 1 (Rs-AFP1) is a 51 amino acid residue plant defensin isolated from radish (Raphanus sativus L.) seeds. The three-dimensional structure in aqueous solution has been determined from two-dimensional super(1)H NMR data recorded at 500 MHz using the DIANA/REDAC calculation protocols. Experimental constraints consisted of 787 interproton distances extracted from NOE cross-peaks, 89 torsional constraints from 106 vicinal interproton coupling constants and 32 stereospecific assignments of prochiral protons. Further refinement by simulated annealing resulted in a set of 20 structures having pairwise root-mean-square differences of 1.35( plus or minus 0.35) Angstrom over the backbone heavy atoms and 2.11( plus or minus 0.46) Angstrom over all heavy atoms. The molecule adopts a compact globular fold comprising an alpha -helix from Asn18 till Leu28 and a triple-stranded beta -sheet ( beta 1 = Lys2-Arg6, beta 2 = His33-Tyr38 and beta 3 = His43-Pro50). The central strand of this beta -sheet is connected by two disulfide bridges (Cys21-Cys45 and Cys25-Cys47) to the alpha -helix. The connection between beta -strand 2 and 3 is formed by a type VIa beta -turn. Even the loop (Pro7 to Asn17) between beta -strand 1 and the alpha -helix is relatively well defined. The structure of Raphanus sativus Antifungal Protein 1 features all the characteristics of the "cysteine stabilized alpha beta motif". A comparison of the complete structure and of the regions important for interaction with the fungal receptor according to a mutational study, is made with the structure of gamma -thionin, a plant defensin that has no antifungal activity. It is concluded that this interaction is both electrostatic and specific, and some possible scenarios for the mode of action are given.
ISSN:0022-2836