Structural insights into the molecular mechanism of phytoplasma immunodominant membrane protein.

Immunodominant membrane protein (IMP) is a prevalent membrane protein in phytoplasma and has been confirmed to be an F-actin-binding protein. However, the intricate molecular mechanisms that govern the function of IMP require further elucidation. In this study, the X-ray crystallographic structure of IMP was determined and insights into its interaction with plant actin are provided. A comparative analysis with other proteins demonstrates that IMP shares structural homology with talin rod domain-containing protein 1 (TLNRD1), which also functions as an F-actin-binding protein. Subsequent molecular-docking studies of IMP and F-actin reveal that they possess complementary surfaces, suggesting a stable interaction. The low potential energy and high confidence score of the IMP-F-actin binding model indicate stable binding. Additionally, by employing immunoprecipitation and mass spectrometry, it was discovered that IMP serves as an interaction partner for the phytoplasmal effector causing phyllody 1 (PHYL1). It was then shown that both IMP and PHYL1 are highly expressed in the S2 stage of peanut witches' broom phytoplasma-infected Catharanthus roseus. The association between IMP and PHYL1 is substantiated through in vivo immunoprecipitation, an in vitro cross-linking assay and molecular-docking analysis. Collectively, these findings expand the current understanding of IMP interactions and enhance the comprehension of the interaction of IMP with plant F-actin. They also unveil a novel interaction pathway that may influence phytoplasma pathogenicity and host plant responses related to PHYL1. This discovery could pave the way for the development of new strategies to overcome phytoplasma-related plant diseases.

Structural and functional analyses of viral H2 protein of the vaccinia virus entry fusion complex.

IMPORTANCE: Vaccinia virus infection requires virus-cell membrane fusion to complete entry during endocytosis; however, it contains a large viral fusion protein complex of 11 viral proteins that share no structure or sequence homology to all the known viral fusion proteins, including type I, II, and III fusion proteins. It is thus very challenging to investigate how the vaccinia fusion complex works to trigger membrane fusion with host cells. In this study, we crystallized the ectodomain of vaccinia H2 protein, one component of the viral fusion complex. Furthermore, we performed a series of mutational, biochemical, and molecular analyses and identified two surface loops containing 170LGYSG174 and 125RRGTGDAW132 as the A28-binding region. We also showed that residues in the N-terminal helical region (amino acids 51-90) are also important for H2 function.

A bacterial binary toxin system that kills both insects and aquatic crustaceans: Photorhabdus insect-related toxins A and B.

Photorhabdus insect-related toxins A and B (PirA and PirB) were first recognized as insecticidal toxins from Photorhabdus luminescens. However, subsequent studies showed that their homologs from Vibrio parahaemolyticus also play critical roles in the pathogenesis of acute hepatopancreatic necrosis disease (AHPND) in shrimps. Based on the structural features of the PirA/PirB toxins, it was suggested that they might function in the same way as a Bacillus thuringiensis Cry pore-forming toxin. However, unlike Cry toxins, studies on the PirA/PirB toxins are still scarce, and their cytotoxic mechanism remains to be clarified. In this review, based on our studies of V. parahaemolyticus PirAvp/PirBvp, we summarize the current understanding of the gene locations, expression control, activation, and cytotoxic mechanism of this type of toxin. Given the important role these toxins play in aquatic disease and their potential use in pest control applications, we also suggest further topics for research. We hope the information presented here will be helpful for future PirA/PirB studies.

Multiple Nucleocapsid Structural Forms of Shrimp White Spot Syndrome Virus Suggests a Novel Viral Morphogenetic Pathway.

White spot syndrome virus (WSSV) is a very large dsDNA virus. The accepted shape of the WSSV virion has been as ellipsoidal, with a tail-like extension. However, due to the scarcity of reliable references, the pathogenesis and morphogenesis of WSSV are not well understood. Here, we used transmission electron microscopy (TEM) and cryogenic electron microscopy (Cryo-EM) to address some knowledge gaps. We concluded that mature WSSV virions with a stout oval-like shape do not have tail-like extensions. Furthermore, there were two distinct ends in WSSV nucleocapsids: a portal cap and a closed base. A C14 symmetric structure of the WSSV nucleocapsid was also proposed, according to our Cryo-EM map. Immunoelectron microscopy (IEM) revealed that VP664 proteins, the main components of the 14 assembly units, form a ring-like architecture. Moreover, WSSV nucleocapsids were also observed to undergo unique helical dissociation. Based on these new results, we propose a novel morphogenetic pathway of WSSV.

An undefined cystatin CsCPI1 from tea plant Camellia sinensis harbors antithrombotic activity.

Tea consumption has been linked to a decreased risk of cardiovascular disease (CVD) mortality, which imposes a heavy burden on the healthcare system; however, which components in tea cause this beneficial effect is not fully understood. Here we uncovered a cystatin (namely CsCPI1), which is a cysteine proteinase inhibitor (CPI) of the tea plant (Camellia sinensis) that promotes antithrombotic activity. Since thrombosis is a common pathogenesis of fatal CVDs, we investigated the effects of CsCPI1, which showed good therapeutic effects in mouse models of thrombotic disease and ischemic stroke. CsCPI1 significantly increases endothelial cell production of nitric oxide (NO) and inhibits platelet aggregation. Notably, CsCPI1 exhibited no cytotoxicity or resistance to pH and temperature changes, which indicates that CsCPI1 might be a potent antithrombotic agent that contributes to the therapeutic effects of tea consumption against CVD. Specifically, the antithrombotic effects of CsCPI1 are distinct from the classical function of plant cystatins against herbivorous insects. Therefore, our study proposes a new potential role of cystatins in CVD prevention and treatment, which requires further study.

Expression of the AHPND Toxins PirAvp and PirBvp Is Regulated by Components of the Vibrio parahaemolyticus Quorum Sensing (QS) System.

Acute hepatopancreatic necrosis disease (AHPND) in shrimp is caused by Vibrio strains that harbor a pVA1-like plasmid containing the pirA and pirB genes. It is also known that the production of the PirA and PirB proteins, which are the key factors that drive the observed symptoms of AHPND, can be influenced by environmental conditions and that this leads to changes in the virulence of the bacteria. However, to our knowledge, the mechanisms involved in regulating the expression of the pirA/pirB genes have not previously been investigated. In this study, we show that in the AHPND-causing Vibrio parahaemolyticus 3HP strain, the pirAvp and pirBvp genes are highly expressed in the early log phase of the growth curve. Subsequently, the expression of the PirAvp and PirBvp proteins continues throughout the log phase. When we compared mutant strains with a deletion or substitution in two of the quorum sensing (QS) master regulators, luxO and/or opaR (luxOD47E, ΔopaR, ΔluxO, and ΔopaRΔluxO), our results suggested that expression of the pirAvp and pirBvp genes was related to the QS system, with luxO acting as a negative regulator of pirAvp and pirBvp without any mediation by opaRvp. In the promoter region of the pirAvp/pirBvp operon, we also identified a putative consensus binding site for the QS transcriptional regulator AphB. Real-time PCR further showed that aphBvp was negatively controlled by LuxOvp, and that its expression paralleled the expression patterns of pirAvp and pirBvp. An electrophoretic mobility shift assay (EMSA) showed that AphBvp could bind to this predicted region, even though another QS transcriptional regulator, AphAvp, could not. Taken together, these findings suggest that the QS system may regulate pirAvp/pirBvp expression through AphBvp.

Novel Algorithm for Improved Protein Classification Using Graph Similarity.

Considerable sequence data are produced in genome annotation projects that relate to molecular levels, structural similarities, and molecular and biological functions. In structural genomics, the most essential task involves resolving protein structures efficiently with hardware or software, understanding these structures, and assigning their biological functions. Understanding the characteristics and functions of proteins enables the exploration of the molecular mechanisms of life. In this paper, we examine the problems of protein classification. Because they perform similar biological functions, proteins in the same family usually share similar structural characteristics. We employed this premise in designing a classification algorithm. In this algorithm, auxiliary graphs are used to represent proteins, with every amino acid in a protein to a vertex in a graph. Moreover, the links between amino acids correspond to the edges between the vertices. The proposed algorithm classifies proteins according to the similarities in their graphical structures. The proposed algorithm is efficient and accurate in distinguishing proteins from different families and outperformed related algorithms experimentally.

Investigating the Viral Suppressor HC-Pro Inhibiting Small RNA Methylation through Functional Comparison of HEN1 in Angiosperm and Bryophyte.

In plants, HEN1-facilitated methylation at 3' end ribose is a critical step of small-RNA (sRNA) biogenesis. A mutant of well-studied Arabidopsis HEN1 (AtHEN1), hen1-1, showed a defective developmental phenotype, indicating the importance of sRNA methylation. Moreover, Marchantia polymorpha has been identified to have a HEN1 ortholog gene (MpHEN1); however, its function remained unfathomed. Our in vivo and in vitro data have shown MpHEN1 activity being comparable with AtHEN1, and their substrate specificity towards duplex microRNA (miRNA) remained consistent. Furthermore, the phylogenetic tree and multiple alignment highlighted the conserved molecular evolution of the HEN1 family in plants. The P1/HC-Pro of the turnip mosaic virus (TuMV) is a known RNA silencing suppressor and inhibits HEN1 methylation of sRNAs. Here, we report that the HC-Pro physically binds with AtHEN1 through FRNK motif, inhibiting HEN1's methylation activity. Moreover, the in vitro EMSA data indicates GST-HC-Pro of TuMV lacks sRNA duplex-binding ability. Surprisingly, the HC-Pro also inhibits MpHEN1 activity in a dosage-dependent manner, suggesting the possibility of interaction between HC-Pro and MpHEN1 as well. Further investigations on understanding interaction mechanisms of HEN1 and various HC-Pros can advance the knowledge of viral suppressors.

Synthesis and biological evaluation of phenothiazine derivative-containing hydroxamic acids as potent class II histone deacetylase inhibitors.

The pathogenesis of Alzheimer's disease (AD) has been associated with dysregulation of histone deacetylases (HDACs). Previously, acridine-based HDAC inhibitors have shown potential in ameliorating HDAC activity and enhancing neurite outgrowth. In this study, the acridine ring was modified using various phenothiazine derivatives. Several resulting compounds exhibited potent enzyme-inhibiting activity towards class II HDACs when compared to the clinically approved HDAC inhibitor SAHA. Compound 4f demonstrated the highest class II HDAC inhibition (IC50 = 4.6-600 nM), as well as promotion of neurite outgrowth. Importantly, compound 4f displayed no cytotoxicity against neuron cells. Compound 4f was further evaluated for cellular effects. Altogether, these findings show a potential strategy in HDAC inhibition for treatment of the neurological disease.

Identification and characterization of l-amino acid oxidase 2 gene in orange-spotted grouper (Epinephelus coioides).

Recently, l-amino acid oxidases (LAAOs) have been identified in several fish species as first-line defense molecules against bacterial infection. Here, we report the cloning and characterization of a fish LAAO gene, EcLAAO2, from orange-spotted grouper (Epinephelus coioides). The full-length cDNA is 3030 bp, with an ORF encoding a protein of 511 amino acids. EcLAAO2 is mainly expressed in the fin, gill, and intestine. Its expression is upregulated in several immune organs after challenge with lipopolysaccharide (LPS) and poly (I:C). The recombinant EcLAAO2 protein (rEcLAAO2), expressed and purified from a baculovirus expression system, was determined to be a glycosylated dimer. According to a hydrogen peroxide-production assay, the recombinant protein was identified as having LAAO enzyme activity with substrate preference for L-Phe and L-Trp, but not L-Lys as other known fish LAAOs. rEcLAAO2 could effectively inhibit the growth of Vibrio parahaemolyticus, Staphylococcus aureus, and Bacillus subtilis while exhibiting less effective inhibition of the growth of Escherichia coli. Finally, protein models based on sequence homology were constructed to predict the three-dimensional structure of EcLAAO2 as well as to explain the difference in substrate specificity between EcLAAO2 and other reported fish LAAOs. In conclusion, this study identifies EcLAAO2 as a novel fish LAAO with a substrate preference distinct from other known fish LAAOs and reveals that it may function against invading pathogens.

A Review of the Functional Annotations of Important Genes in the AHPND-Causing pVA1 Plasmid.

Acute hepatopancreatic necrosis disease (AHPND) is a lethal shrimp disease. The pathogenic agent of this disease is a special Vibrio parahaemolyticus strain that contains a pVA1 plasmid. The protein products of two toxin genes in pVA1, pirAvp and pirBvp, targeted the shrimp's hepatopancreatic cells and were identified as the major virulence factors. However, in addition to pirAvp and pirBvp, pVA1 also contains about ~90 other open-reading frames (ORFs), which may encode functional proteins. NCBI BLASTp annotations of the functional roles of 40 pVA1 genes reveal transposases, conjugation factors, and antirestriction proteins that are involved in horizontal gene transfer, plasmid transmission, and maintenance, as well as components of type II and III secretion systems that may facilitate the toxic effects of pVA1-containing Vibrio spp. There is also evidence of a post-segregational killing (PSK) system that would ensure that only pVA1 plasmid-containing bacteria could survive after segregation. Here, in this review, we assess the functional importance of these pVA1 genes and consider those which might be worthy of further study.

Structural insight into the differential interactions between the DNA mimic protein SAUGI and two gamma herpesvirus uracil-DNA glycosylases.

Uracil-DNA glycosylases (UDGs) are conserved DNA-repair enzymes that can be found in many species, including herpesviruses. Since they play crucial roles for efficient viral DNA replication in herpesviruses, they have been considered as potential antiviral targets. In our previous work, Staphylococcus aureus SAUGI was identified as a DNA mimic protein that targets UDGs from S. aureus, human, Herpes simplex virus (HSV) and Epstein-Barr virus (EBV). Interestingly, SAUGI has the strongest inhibitory effects with EBVUDG. Here, we determined complex structures of SAUGI with EBVUDG and another γ-herpesvirus UDG from Kaposi's sarcoma-associated herpesvirus (KSHVUDG), which SAUGI fails to effectively inhibit. Structural analysis of the SAUGI/EBVUDG complex suggests that the additional interaction between SAUGI and the leucine loop may explain why SAUGI shows the highest binding capacity with EBVUDG. In contrast, SAUGI appears to make only partial contacts with the key components responsible for the compression and stabilization of the DNA backbone in the leucine loop extension of KSHVUDG. The findings in this study provide a molecular explanation for the differential inhibitory effects and binding strengths that SAUGI has on these two UDGs, and the structural basis of the differences should be helpful in developing inhibitors that would interfere with viral DNA replication.

A shrimp glycosylase protein, PmENGase, interacts with WSSV envelope protein VP41B and is involved in WSSV pathogenesis.

Viral glycoproteins are expressed by many viruses, and during infection they usually play very important roles, such as receptor attachment or membrane fusion. The mature virion of the white spot syndrome virus (WSSV) is unusual in that it contains no glycosylated proteins, and there are currently no reports of any glycosylation mechanisms in the pathogenesis of this virus. In this study, we cloned a glycosylase, mannosyl-glycoprotein endo-ß-N-acetylglucosaminidase (ENGase, EC 3.2.1.96), from Penaeus monodon and found that it was significantly up-regulated in WSSV-infected shrimp. A yeast two-hybrid assay showed that PmENGase interacted with both structural and non-structural proteins, and GST-pull down and co-immunoprecipitation (Co-IP) assays confirmed its interaction with the envelope protein VP41B. In the WSSV challenge tests, the cumulative mortality and viral copy number were significantly decreased in the PmEngase-silenced shrimp, from which we conclude that shrimp glycosylase interacts with WSSV in a way that benefits the virus. Lastly, we speculate that the deglycosylation activity of PmENGase might account for the absence of glycosylated proteins in the WSSV virion.

Structural insights into the interaction between phytoplasmal effector causing phyllody 1 and MADS transcription factors.

Phytoplasmas are bacterial plant pathogens which can induce severe symptoms including dwarfism, phyllody and virescence in an infected plant. Because phytoplasmas infect many important crops such as peanut and papaya they have caused serious agricultural losses. The phytoplasmal effector causing phyllody 1 (PHYL1) is an important phytoplasmal pathogenic factor which affects the biological function of MADS transcription factors by interacting with their K (keratin-like) domain, thus resulting in abnormal plant developments such as phyllody. Until now, lack of information on the structure of PHYL1 has prevented a detailed understanding of the binding mechanism between PHYL1 and the MADS transcription factors. Here, we present the crystal structure of PHYL1 from peanut witches'-broom phytoplasma (PHYL1PnWB ). This protein was found to fold into a unique α-helical hairpin with exposed hydrophobic residues on its surface that may play an important role in its biological function. Using proteomics approaches, we propose a binding mode of PHYL1PnWB with the K domain of the MADS transcription factor SEPALLATA3 (SEP3_K) and identify the residues of PHYL1PnWB that are important for this interaction. Furthermore, using surface plasmon resonance we measure the binding strength of PHYL1PnWB proteins to SEP3_K. Lastly, based on confocal images, we found that α-helix 2 of PHYL1PnWB plays an important role in PHYL1-mediated degradation of SEP3. Taken together, these results provide a structural understanding of the specific binding mechanism between PHYL1PnWB and SEP3_K.

Vaccinia viral A26 protein is a fusion suppressor of mature virus and triggers membrane fusion through conformational change at low pH.

Vaccinia mature virus requires A26 envelope protein to mediate acid-dependent endocytosis into HeLa cells in which we hypothesized that A26 protein functions as an acid-sensitive membrane fusion suppressor. Here, we provide evidence showing that N-terminal domain (aa1-75) of A26 protein is an acid-sensitive region that regulates membrane fusion. Crystal structure of A26 protein revealed that His48 and His53 are in close contact with Lys47, Arg57, His314 and Arg312, suggesting that at low pH these His-cation pairs could initiate conformational changes through protonation of His48 and His53 and subsequent electrostatic repulsion. All the A26 mutant mature viruses that interrupted His-cation pair interactions of His48 and His 53 indeed have lost virion infectivity. Isolation of revertant viruses revealed that second site mutations caused frame shifts and premature termination of A26 protein such that reverent viruses regained cell entry through plasma membrane fusion. Together, we conclude that viral A26 protein functions as an acid-sensitive fusion suppressor during vaccinia mature virus endocytosis.

Structural Insights to the Heterotetrameric Interaction between the Vibrio parahaemolyticus PirAvp and PirBvp Toxins and Activation of the Cry-Like Pore-Forming Domain.

Acute hepatopancreatic necrosis disease (AHPND) is a newly emergent penaeid shrimp disease which can cause 70-100% mortality in Penaeus vannamei and Penaeus monodon, and has resulted in enormous economic losses since its appearance. AHPND is caused by the specific strains of Vibrio parahaemolyticus that harbor the pVA1 plasmid and express PirAvp and PirBvp toxins. These two toxins have been reported to form a binary complex. When both are present, they lead to the death of shrimp epithelial cells in the hepatopancreas and cause the typical histological symptoms of AHPND. However, the binding mode of PirAvp and PirBvp has not yet been determined. Here, we used isothermal titration calorimetry (ITC) to measure the binding affinity of PirAvp and PirBvp. Since the dissociation constant (Kd = 7.33 ± 1.20 µM) was considered too low to form a sufficiently stable complex for X-ray crystallographic analysis, we used alternative methods to investigate PirAvp-PirBvp interaction, first by using gel filtration to evaluate the molecular weight of the PirAvp/PirBvp complex, and then by using cross-linking and hydrogen-deuterium exchange (HDX) mass spectrometry to further understand the interaction interface between PirAvp and PirBvp. Based on these results, we propose a heterotetrameric interaction model of this binary toxin complex. This model provides insight of how conformational changes might activate the PirBvp N-terminal pore-forming domain and should be helpful for devising effective anti-AHPND strategies in the future.

New paradigm of functional regulation by DNA mimic proteins: Recent updates.

For many, "DNA mimic protein" (DMP) remains an unfamiliar term. The key feature of these proteins is their DNA-like shape and charge distribution, and they affect the activity of DNA-binding proteins by occupying their DNA-binding domains. Functionally, DMPs regulate mechanisms such as gene expression, restriction, and DNA repair as well as the nucleosome package. Although a few DMPs, such as phage uracil DNA glycosylase inhibitor (UGI) and overcome classical restriction (Ocr), were reported about 20 years ago, only a small number of DMPs have been studied to date. In 2014, we reviewed the functional and structural features of 16 DMPs that were known at the time. Now, seven new DMPs, namely anti-CRISPR suppressors AcrF2, AcrF10 and AcrIIA4, human immunodeficiency virus essential factor VPR, multi-functional inhibitor anti-restriction nuclease (Arn), translational regulator AbbA, and putative Z-DNA mimic MBD3, have been reported. In addition, further study of two previously known DMPs, DMP19 and SAUGI, increased our knowledge of their importance and function. Here, we discuss these updated results and address how several characteristics of the structure/sequence of DMPs (e.g. the DNA-like charge distribution and structural D/E-rich repeats) might someday be used to identify new DMPs using bioinformatic approach. © 2018 IUBMB Life, 71(5):539-548, 2019.

Gene-to-Gene Network Analysis of the Mediation of Plant Innate Immunity by the Eliciting Plant Response-Like 1 (Epl1) Elicitor of Trichoderma formosa.

A new clade, Trichoderma formosa, secretes eliciting plant response-like 1 (Epl1), a small peptide elicitor that stimulates plant immunity. Nicotiana benthamiana pretreated with Epl1 for 3 days developed immunity against Tomato mosaic virus (ToMV) infection. The transcriptome profiles of T. formosa and N. benthamiana were obtained by deep sequencing; the transcript of Epl1 is 736 nt in length and encodes a 12-kDa peptide. Identifying critical genes in Epl1-mediated immunity was challenging due to high similarity between the transcriptome expression profiles of Epl1-treated and ToMV-infected N. benthamiana samples. Therefore, an efficient bioinformatics data mining approach was used for high-throughput transcriptomic assays in this study. We integrated gene-to-gene network analysis into the ContigViews transcriptome database, and genes related to jasmonic acid and ethylene signaling, salicylic acid signaling, leucine-rich repeats, transcription factors, and histone variants were hubs in the gene-to-gene networks. In this study, the Epl1 of T. formosa triggers plant immunity against various pathogen infections. Moreover, we demonstrated that high-throughput data mining and gene-to-gene network analysis can be used to identify critical candidate genes for further studies on the mechanisms of plant immunity.

Dual Inhibition of PIK3C3 and FGFR as a New Therapeutic Approach to Treat Bladder Cancer.

Purpose: MPT0L145 has been developed as a FGFR inhibitor exhibiting significant anti-bladder cancer activity in vitro and in vivo via promoting autophagy-dependent cell death. Here, we aim to elucidate the underlying mechanisms.Experimental Design: Autophagy flux, morphology, and intracellular organelles were evaluated by Western blotting, transmission electron microscope, and fluorescence microscope. Molecular docking and surface plasmon resonance assay were performed to identify drug-protein interaction. Lentiviral delivery of cDNA or shRNA and CRISPR/Cas9-mediated genome editing was used to modulate gene expression. Mitochondrial oxygen consumption rate was measured by a Seahorse XFe24 extracellular flux analyzer, and ROS level was measured by flow cytometry.Results: MPT0L145 persistently increased incomplete autophagy and phase-lucent vacuoles at the perinuclear region, which were identified as enlarged and alkalinized late-endosomes. Screening of a panel of lipid kinases revealed that MPT0L145 strongly inhibits PIK3C3 with a Kd value of 0.53 nmol/L. Ectopic expression of PIK3C3 reversed MPT0L145-increased cell death and incomplete autophagy. Four residues (Y670, F684, I760, D761) at the ATP-binding site of PIK3C3 are important for the binding of MPT0L145. In addition, MPT0L145 promotes mitochondrial dysfunction, ROS production, and DNA damage, which may in part, contribute to cell death. ATG5-knockout rescued MPT0L145-induced cell death, suggesting simultaneous induction of autophagy is crucial to its anticancer activity. Finally, our data demonstrated that MPT0L145 is able to overcome cisplatin resistance in bladder cancer cells.Conclusions: MPT0L145 is a first-in-class PIK3C3/FGFR inhibitor, providing an innovative strategy to design new compounds that increase autophagy, but simultaneously perturb its process to promote bladder cancer cell death. Clin Cancer Res; 24(5); 1176-89. ©2017 AACR.

Structural Insights into the Cytotoxic Mechanism of Vibrio parahaemolyticus PirAvp and PirBvp Toxins.

In aquaculture, shrimp farming is a popular field. The benefits of shrimp farming include a relatively short grow-out time, high sale price, and good cost recovery. However, outbreaks of serious diseases inflict serious losses, and acute hepatopancreatic necrosis disease (AHPND) is an emerging challenge to this industry. In South American white shrimp (Penaeus vannamei) and grass shrimp (Penaeus monodon), this disease has a 70-100% mortality. The pathogenic agent of AHPND is a specific strain of Vibrio parahaemolyticus which contains PirAvp and PirBvp toxins encoded in the pVA1 plasmid. PirAvp and PirBvp have been shown to cause the typical histological symptoms of AHPND in infected shrimps, and in this review, we will focus on our structural understanding of these toxins. By analyzing their structures, a possible cytotoxic mechanism, as well as strategies for anti-AHPND drug design, is proposed.