A novel anti-lipopolysaccharide factor Sp ALF6 in mud crab Scylla paramamosain exhibiting different antimicrobial activity from its single amino acid mutant

Hou, Zhiguo; Wang, Yuan; Hui, Kai-Min; Fang, Wenhong; Zhang, Jingxiao; Ma, Hongyu; Li, Xin-Cang

doi:10.1016/j.dci.2017.02.009

Cited by 35 publications

(14 citation statements)

References 48 publications

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“…Enzyme-linked immunosorbent assay was conducted to evaluate the binding ability of NOD1-LRR to ultrapure LPS-B5, ultrapure LPS-EK, Synthetic Lipid A, iE-DAP (γ- d -Glu-mDAP, tlrl-dap, Invivogen), and β-1,3-Glucan (89862, Sigma). Each well of a microtiter plate was coated with 100 µl of 20 µg/ml ultrapure LPS, Lipid A, iE-DAP, or β-1,3-Glucan, and then incubated overnight at 37°C following a previously described protocol ( 35 ). Wells incubated with 100 µl of 50 mM Tris–HCl were used as negative control.…”

Section: Methodsmentioning

confidence: 99%

Recognition of Lipopolysaccharide and Activation of NF-κB by Cytosolic Sensor NOD1 in Teleost Fish

Wang

Gao

et al. 2018

Front. Immunol.

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Lipopolysaccharide (LPS) is the major component of the outer membrane of Gram-negative bacteria. This molecule can induce strong immune response and various biological effects. In mammals, TLR4 can recognize LPS and induce inflammatory response. However, the innate receptor in fish for recognizing LPS remains ambiguous. LPS can invade the cytoplasm via outer membrane vesicles produced by Gram-negative bacteria and could be detected by intracellular receptor caspase-11 in mammals, so, there may also exist the intracellular receptors that can recognize LPS in fish. NOD1 is a member of NOD-like receptors family and can recognize the iE-DAP in the cytoplasm in mammals. In fish, NOD1 can also respond to infection of Gram-negative bacteria and may play an important role in the identification of bacterial components. In this study, to study whether NOD1 is a recognition receptor for LPS, we detected the expression of NOD1 and several cytokines at transcript levels to determine whether LPS can induce inflammatory response in teleost fish and NOD1 can respond to LPS. Then, we perform the binding analysis between NOD1 and ultrapure LPS by using Streptavidin pulldown assay and enzyme-linked immunosorbent assay to prove that NOD1 can be combined with LPS, and using dual luciferase reporter gene assay to verify the signal pathways activated by NOD1. Next, through cell viability analysis, we proved that LPS-induced cytotoxicity can be mediated by NOD1 in fish. The results showed that NOD1 can identify LPS and activate the NF-κB signal pathway by recruiting RIPK2 and then promoting the expression of inflammatory cytokines to induce the resistance of organism against bacterial infection.

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Section: Methodsmentioning

confidence: 99%

Recognition of Lipopolysaccharide and Activation of NF-κB by Cytosolic Sensor NOD1 in Teleost Fish

Wang

Gao

et al. 2018

Front. Immunol.

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“…The recombinant ALF proteins exhibited in vitro antimicrobial activities (8) and pre-injection of recombinant ALF protein could reduce cumulative mortality of shrimp caused by Vibrio harveyi infection (9). In mud crab Scylla paramamosain , a single amino acid mutant of SpALF6 obviously changed its antimicrobial activities (10). These data revealed that ALFs were essential effectors in innate immune system of crustacean.…”

Section: Introductionmentioning

confidence: 99%

Characterization of a Lymphoid Organ Specific Anti-lipopolysaccharide Factor From Shrimp Reveals Structure-Activity Relationship of the LPS-Binding Domain

et al. 2019

Front. Immunol.

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Anti-lipopolysaccharide factor (ALF) is a kind of important antimicrobial peptides with broad-spectrum antimicrobial activities. The LPS-binding domain (LBD) contributes to the major antimicrobial activity of ALF. However, LBDs from different ALFs share low sequence similarity. The general character of LBDs needs to be elucidated to understand the molecular mechanism of their function and facilitate LBD-original drug design. Here we identified a lymphoid organ specifically expressed ALF , designated as FcALF8 , from the Chinese shrimp Fenneropenaeus chinensis . The synthetic LBD peptide of FcALF8 (LBD8) showed strong antibacterial activities to the pathogenic Vibrio , such as Vibrio alginolyticus, Vibrio harveyi , and Photobacterium damselae with a MIC value of 0.5–1, 1–2, and 1–2 μM, respectively. FcALF8 knock-down using dsRNA led to significant increase of the viable bacteria in the lymphoid organ and hepatopancreas of shrimp upon V. harveyi infection. On the contrary, the proliferation of V. harveyi in the shrimp lymphoid organ and hepatopancreas significantly decreased after infected by LBD8 pre-incubated V. harveyi . Sequence alignments showed that the LBDs from 39 ALFs shared only two identical cysteine residues. However, 17 of the total 22 LBD residues showed high similarity when the amino acids were classified into hydrophobic and hydrophilic ones. A further activity analysis on modified LBD8 peptides showed that the antibacterial activity of LBD8 was lost after linearization and apparently weakened after changing the amino acid property at certain positions. The data indicated that the disulfide bond and amino acid property contributed to the conservation of the functional domain. To the best of our knowledge, this is the first identified ALFs specifically expressed in the lymphoid organ of shrimp with strong antibacterial activity. The present data will give creative instructions for the design of LBD-originated antimicrobial agents.

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“…RALFPm3 could bind to lipid A of LPS [ 46 ]. Recently, a novel ALF named SpALF6, which was isolated from the mud crab Scylla paramamosain , and its variant SpALF6-V (H46R and A110P) possessed strong binding activity to LPS and exhibited broad antimicrobial activity against Gram-negative bacteria (such as Vibrio and E. coli ), Gram-positive bacteria (such as S. aureus ) and fungi (such as C. albicans ) [ 47 ].…”

Section: Antibacterial/antiendotoxic Peptides or Proteins From Varmentioning

confidence: 99%

“…Pv5 derivative-Pt5e was also successfully expressed in E. coli and displayed dual antibacterial and LPS neutralizing function in vitro and in vivo [ 60 ]. The expressed SpALF6 in E. coli had potent antibacterial and antifungal activity and also recognized LPS [ 47 ].…”

Section: Heterologous Expression Of Antibacterial and Antiendotoximentioning

confidence: 99%

“…LPS is then recognized by toll-like receptor 4 (TRL4) on the surface of the phagocytic cells, which in turn activates the MAPK and NF-κB pathways, which can induce the production of pro-inflammatory cytokines. The mechanism of peptides or proteins in neutralizing LPS and inhibiting the inflammatory response can be divided into (i) a direct effect: many bifunctional peptides (such as Hc-CATH) or proteins (such as SpALF6) can directly bind with LPS, which thus inhibits the binding of LPS to the TLR4/MD2 complex and the activation of inflammatory response pathways [ 18 , 47 ]; and (ii) an indirect effect: peptides (such as LL37 and defensin) or proteins competitively bind to CD14 or TLR4, thereby indirectly inhibiting LPS-induced reactions downstream ( Figure 1 ) [ 82 , 93 ].…”

Section: Mechanism Of Marine Peptides or Proteins In Neutralizing mentioning

confidence: 99%

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Recent Advances in Antibacterial and Antiendotoxic Peptides or Proteins from Marine Resources

Wang

2018

Marine Drugs

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Infectious diseases caused by Gram-negative bacteria and sepsis induced by lipopolysaccharide (LPS) pose a major threat to humans and animals and cause millions of deaths each year. Marine organisms are a valuable resource library of bioactive products with huge medicinal potential. Among them, antibacterial and antiendotoxic peptides or proteins, which are composed of metabolically tolerable residues, are present in many marine species, including marine vertebrates, invertebrates and microorganisms. A lot of studies have reported that these marine peptides and proteins or their derivatives exhibit potent antibacterial activity and antiendotoxic activity in vitro and in vivo. However, their categories, heterologous expression in microorganisms, physicochemical factors affecting peptide or protein interactions with bacterial LPS and LPS-neutralizing mechanism are not well known. In this review, we highlight the characteristics and anti-infective activity of bifunctional peptides or proteins from marine resources as well as the challenges and strategies for further study.

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A novel anti-lipopolysaccharide factor Sp ALF6 in mud crab Scylla paramamosain exhibiting different antimicrobial activity from its single amino acid mutant

Cited by 35 publications

References 48 publications

Recognition of Lipopolysaccharide and Activation of NF-κB by Cytosolic Sensor NOD1 in Teleost Fish

Recognition of Lipopolysaccharide and Activation of NF-κB by Cytosolic Sensor NOD1 in Teleost Fish

Characterization of a Lymphoid Organ Specific Anti-lipopolysaccharide Factor From Shrimp Reveals Structure-Activity Relationship of the LPS-Binding Domain

Recent Advances in Antibacterial and Antiendotoxic Peptides or Proteins from Marine Resources

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