The major nectar protein of Brassica rapa is a non-specific lipid transfer protein, BrLTP2.1, with strong antifungal activity

Abstract

Nectar is one of the key rewards mediating plant–mutualist interactions. In addition to sugars, nectars often contain many other compounds with important biological functions, including proteins. This study was undertaken to assess the proteinaceous content of Brassica rapa nectar. SDS-PAGE analysis of raw B. rapa nectar revealed the presence of ~10 proteins, with a major band at ~10 kDa. This major band was found to contain a non-specific lipid transfer protein encoded by B. rapa locus Bra028980 and subsequently termed BrLTP2.1. Sequence analysis of BrLTP2.1 predicted the presence of a signal peptide required for secretion from the cell, eight cysteines, and a mature molecular mass of 7.3 kDa. Constitutively expressed BrLTP2.1–GFP in Arabidopsis displayed accumulation patterns consistent with secretion from nectary cells. BrLTP2.1 was also found to have relatively high sequence similarity to non-specific lipid-transfer proteins with known functions in plant defense, including Arabidopsis DIR1. Heterologously expressed and purified BrLTP2.1 was extremely heat stable and bound strongly to saturated free fatty acids, but not methyl jasmonate. Recombinant BrLTP2.1 also had direct antimicrobial activity against an extensive range of plant pathogens, being particularly effective against necrotrophic fungi. Taken together, these results suggest that BrLTP2.1 may function to prevent microbial growth in nectars.

Structural characterization and in vitro lipid binding studies of non-specific lipid transfer protein 1 (nsLTP1) from fennel (Foeniculum vulgare) seeds

“…7). As supported by previous lipid binding studies, saturated FAs with relatively shorter chain length allow a limited number of hydrophobic interactions resulting in relatively lower binding efficiency as compared to saturated FAs with longer chain length 47,56,57 . On the other hand, significantly higher binding efficiency (***P < 0.001) was noted for linoleic acid, followed by linolenic acid (Fig.…”

“…7). The presence of one cis-double bond in FAs with a chain length of C18 (as in linoleic acid) has a higher affinity for the protein as it enforces the FA to a curved shape for easier accommodation in the binding site 23,[56][57][58] . Moreover, the binding assay revealed an initial increase in the percentage of fennel nsLTP1-TNS fluorescence at the lower concentration of displacing FAs, followed by a decrease in the intensity at higher FA concentration (Fig.…”

Proteomics and post-secretory content adjustment of Nicotiana tabacum nectar

“…Some isolated nectarins, e.g. a non-specific lipid transfer protein, showed strong and direct antifungal activity (Schmitt et al 2018).…”

The role of alanine synthesis and nitrate‐induced nitric oxide production during hypoxia stress in Cucurbita pepo nectaries

“…Nectar solutes are typically dominated by sugars, but can also contain various compounds, including proteins (Carter et al ., 1999; Schmitt et al ., 2018), oils (Dötterl and Schäffler, 2007), alkaloids (Deinzer et al ., 1977) and other specialized metabolites (Roy et al ., 2017). Previous studies have reported millimolar concentrations of amino acids in the floral nectars of many species (Baker, 1977), including tobacco (Tiedge and Lohaus, 2017), squash ( Cucurbita pepo ) (Nepi et al ., 2012) and Brassica napus (Bertazzini and Forlani, 2016).…”