Bleogens: Cactus-Derived Anti-Candida Cysteine-Rich Peptides with Three Different Precursor Arrangements.

Shining Loo,Antony Kam,James P Tam,Tianshu Xiao

doi:10.3389/fpls.2017.02162

Abstract

Cysteine-rich peptides (CRPs) play important host-defense roles in plants. However, information concerning CRPs in the Cactaceae (cactus) family is limited, with only a single cactus-derived CRP described to date. Here, we report the identification of 15 novel CRPs with three different precursor architectures, bleogens pB1-15 from Pereskia bleo of the Cactaceae family. By combining proteomic and transcriptomic methods, we showed that the prototype, bleogen pB1, contained 36 amino acid residues, a six-cysteine motif typical of the six-cysteine-hevein-like peptide (6C-HLP) family, and a type I two-domain precursor consisting of an endoplasmic reticulum (ER) and a mature domain. In contrast, the precursors of the other 14 bleogens contained a type II three-domain architecture with a propeptide domain inserted between the ER and the mature bleogen domain. Four of these 14 bleogens display a third type of architecture with a tandemly repeating bleogen domain. A search of the Onekp database revealed that <1% plant species possess three different precursor architectures for the biosynthesis of 6C-HLPs, including Lophophora williamsii, Pereskia aculeate, Portulaca cryptopetala, Portulaca oleracea, Portulaca suffruticosa, and Talinum sp. NMR analysis confirmed that bleogen pB1 has cystine-knot disulfide connectivity as well as a two-beta-sheet and a four-loop structural fold that is similar to other 6C-HLPs. Sequence analysis, structural studies, and in silico modeling revealed that bleogen pB1 has a cation-polar-cation motif, a signature heparin-binding motif that was confirmed by heparin affinity chromatography. Cell-based assays showed that bleogen pB1 is non-toxic to mammalian cells but functions as an anti-Candida peptide. Taken together, our findings provide insight into the occurrence, functions and precursor architectures of CRPs in the cactus family.

Highlights

Pereskia bleo (Kunth) DC, known as rose cactus, is an herb belonging to the Cactaceae family that is commonly used in Southeast Asia
In a mass-spectrometry-driven profiling and discovery program to identify cysteine-rich peptides (CRPs) in medicinal plants, we found a cluster of CRPs ranging from 3 to 5 kDa from aqueous extracts of the medicinal cactus plant Pereskia bleo, belonging to the Cactaceae family
Since bleogen pB1 is one of the major CRP in Pereskia bleo leaf, flower and seed extracts, it was selected for purification and characterization in a scaled-up aqueous extract of Pereskia bleo leaves

Summary

Introduction

Pereskia bleo (Kunth) DC, known as rose cactus, is an herb belonging to the Cactaceae family that is commonly used in Southeast Asia. Small molecule metabolites and proteins represent a major source of drug leads in natural products. Multiple disulfide-constrained peptides derived from medicinal plants represent an underexplored area in drug discovery. These plantderived peptides, the highly disulfide-crosslinked cysteine-rich peptides (CRPs) that have three to five disulfide bonds and a molecular weight between 2 and 6 kDa, are hyperstable, and could provide a potential source of leads for drug development in the neglected chemical space between small molecule metabolites and proteins (Nguyen et al, 2013). Many CRPs are known to be plant defense molecules, which act as antimicrobials, insecticides (Broekaert et al, 1995, 1997; Silverstein et al, 2007; Odintsova and Egorov, 2012; Nawrot et al, 2014), proteinase inhibitors (Hamato et al, 1995; Nguyen et al, 2014, 2015a; Loo et al, 2016), or immunestimulating agents (Nguyen et al, 2016)

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Discussion

Conclusion