Structure of SPH (self-incompatibility protein homologue) proteins: a widespread family of small, highly stable, secreted proteins.

Karthik V Rajasekar,Rachel J Coulthard,Eva I Hyde,Gillian L Reynolds,Michael J Wheeler,Jon P Ride,Shuangxi Ji,Lorna J Smith,Peter J Winn

doi:10.1042/bcj20180828

Karthik V Rajasekar, Rachel J Coulthard + Show 7 more

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https://doi.org/10.1042/bcj20180828

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Abstract

SPH (self-incompatibility protein homologue) proteins are a large family of small, disulfide-bonded, secreted proteins, initially found in the self-incompatibility response in the field poppy (Papaver rhoeas), but now known to be widely distributed in plants, many containing multiple members of this protein family. Using the Origami strain of Escherichia coli, we expressed one member of this family, SPH15 from Arabidopsis thaliana, as a folded thioredoxin fusion protein and purified it from the cytosol. The fusion protein was cleaved and characterised by analytical ultracentrifugation, circular dichroism and nuclear magnetic resonance (NMR) spectroscopy. This showed that SPH15 is monomeric and temperature stable, with a β-sandwich structure. The four strands in each sheet have the same topology as the unrelated proteins: human transthyretin, bacterial TssJ and pneumolysin, with no discernible sequence similarity. The NMR-derived structure was compared with a de novo model, made using a new deep learning algorithm based on co-evolution/correlated mutations, DeepCDPred, validating the method. The DeepCDPred de novo method and homology modelling to SPH15 were then both used to derive models of the 3D structure of the three known PrsS proteins from P. rhoeas, which have only 15-18% sequence homology to SPH15. The DeepCDPred method gave models with lower discreet optimised protein energy scores than the homology models. Three loops at one end of the poppy structures are postulated to interact with their respective pollen receptors to instigate programmed cell death in pollen tubes.

Highlights

Plants express a large number and variety of small, disulfide-bonded, secreted peptides, often acting in defence roles, or in cell signalling
75 members of the SPH family are identified in A. thaliana in the Pfam protein domain database (PF05938) [8], with 120 identified using the bioinformatics programme SPADA [3,8]
SPH proteins and domains have been found in most dicotyledonous plants, with over 1800 homologous sequences identified from 71 species in Pfam [8], and more than 2500 sequences identified in the InterPro protein family database (IPR010264) [10]

Summary

Introduction

Plants express a large number and variety of small, disulfide-bonded, secreted peptides, often acting in defence roles, or in cell signalling. MODELLER [37] was used to model the poppy proteins, based on the structure of SPH15, with two different methods of sequence alignment. The sequence alignment of the SPH domain of SPH15 from A. thaliana and the three PrsS proteins from P. rhoeas is shown, together with the consensus from 759 sequences from HHblits [14] and the secondary structure prediction, showing nine β-strands, from SPIDER2 [16].

Results

Conclusion