Solution structure and lipid binding of a nonspecific lipid transfer protein extracted from maize seeds

Abstract

The three-dimensional solution structure of a nonspecific lipid transfer protein extracted from maize seeds determined by ' H NMR spectroscopy is described. This cationic protein consists of 93 amino acid residues. Its structure was determined from 1,091 NOE-derived distance restraints, including 929 interresidue connectivities and 197 dihedral restraints (4,$, x,) derived from NOES and 3J coupling constants. The global fold involving four helical fragments connected by three loops and a C-terminal tail without regular secondary structures is stabilized by four disulfide bridges. The most striking feature of this structure is the existence of an internal hydrophobic cavity running through the whole molecule. The global fold of this protein, very similar to that of a previously described lipid transfer protein extracted from wheat seeds (Gincel E et al., 1994, Eur J Biochem 226:413-422) constitutes a new architecture for a-class proteins. 'H NMR and fluorescence studies show that this protein forms well-defined complexes in aqueous solution with lysophosphatidylcholine. Dissociation constants, K d , of 1.9 f 0.6 x M and M were obtained with lyso-C,, and -Clz, respectively. A structure model for a lipidprotein complex is proposed in which the aliphatic chain of the phospholipid is inserted in the internal cavity and the polar head interacts with the charged side chains located at one end of this cavity. Our model for the lipidprotein complex is qualitatively very similar to the recently published crystal structure (Shin DH et al., 1995, Structure 3:189-199).

The Nonspecific Lipid Transfer Protein AtLtpI-4 Is Involved in Suberin Formation of Arabidopsis thaliana Crown Galls

“…Hence, shorter fatty acids possibly might bind with lower affinities, whereas longer fatty acids bind tighter, as they completely fill up the hydrophobic chamber and thereby expel all solvent molecules from the pocket. This hypothesis is consistent with data indicating lower affinities for shorter aliphatic chains (Gomar et al, 1996) and structure analyses that show that LTP proteins also can accommodate quite bulky cargo, such as two fatty acids inside rice or wheat LTP1 (Charvolin et al, 1999;Cheng et al, 2004) or a prostaglandin B2 inside wheat nsLTP (TassinMoindrot et al, 2000). Thus, LTP proteins seem to have a highly adaptable cleft (Lerche et al, 1997) that allows them to bind a wide range of hydrophobic molecules, with binding strength being mainly scaled by the number of interactions and restrained by geometric boundaries of the cleft volume (Zachowski et al, 1998).…”

1H NMR and fluorescence studies of the complexation of DMPG by wheat non‐specific lipid transfer protein. Global fold of the complex

“…E-mail : sodano@cnrs-orleans.fr helices and a C-terminal fragment organized around a long internal cavity which has been suggested to be the lipid binding site. This type of organization was then confirmed with the maize ns-LTP structure that we established in solution [3] and in crystal by Shin et al [4] and more recently with the barley ns-LTP structure [5]. The crystal structure of the complex of the maize protein with palmitic acid [4], the solution structure of its complex with l-palmitoyl-2-lysophosphatidylcholine [3] and the barley ns-LTP complex with palmitoylCoA [6] demonstrated the capacity of ns-LTPs to incorporate a long fatty acid chain in their internal cavity.…”

“…This type of organization was then confirmed with the maize ns-LTP structure that we established in solution [3] and in crystal by Shin et al [4] and more recently with the barley ns-LTP structure [5]. The crystal structure of the complex of the maize protein with palmitic acid [4], the solution structure of its complex with l-palmitoyl-2-lysophosphatidylcholine [3] and the barley ns-LTP complex with palmitoylCoA [6] demonstrated the capacity of ns-LTPs to incorporate a long fatty acid chain in their internal cavity. In the present work and in contrast with previous conclusions provided by the study of barley ns-LTP [6], we demonstrate that the wheat ns-LTP can also incorporate 1,2-dimyristoylphosphatidylgrycerol (DMPG) which bears two acyl chains as natural lipid components recovered in cell membranes.…”

“…Titanium particles were removed by filtration on a 0.2 urn Millipore filter. Lipid titration of wheat ns-LTP was carried out by fluorescence spectroscopy as previously described [3]. The dissociation constant (K¿) and the number of binding sites («) were determined by fitting experimental curves with a non-cooperative model assuming independent and identical sites [8].…”

Plant lipid transfer proteins: Evolution, expression and function

“…The crystal structure has been determined for several nsLTPs, among them one from peach, one from rice, one from Arabidopsis and one from maize (Gomar et al, 1996;Poznanski et al, 1999;Lascombe et al, 2008). The structural studies have determined that the folding of nsLTP proteins occurs in two different ways, namely the Type 1 and the Type 2 fold.…”