Dataset Information

Prediction and characterization of a novel hemocyanin-derived antimicrobial peptide from shrimp Litopenaeus vannamei.

ABSTRACT: Hemocyanin, the multifunctional glycoprotein in the hemolymph of invertebrates, can generate various antimicrobial peptides (AMPs). Given the rising interest in the use of natural therapeutic agents such as AMPs, alternative and more efficient methods for their generation are being explored. In this work, free online software was first applied to predict the generation of antimicrobial peptides from the large subunit of Litopenaeus vannamei hemocyanin. Twenty potential antimicrobial peptides ranging from 1.5 to 1.9 kDa were predicted, five of which had ?-helical structures and were selected for antibacterial activity testing. The results indicated that these five peptides had antibacterial activity against seven different bacteria. Of the five peptides, one peptide, designated L1, had the strongest antibacterial activity against both Gram-negative and Gram-positive bacteria. Moreover, CD and NMR data showed that L1 had both ?-helical and ?-turns structural composition, and that these structures were essential for L1's antibacterial activity. Furthermore, SEM analysis revealed that peptide L1 had broad-spectrum activity against both Gram-positive and Gram-negative bacteria, as it could destroy the bacterial cell walls and kill the bacteria. Thus, L1 is a very potent antimicrobial peptide that can be exploited and used in antibacterial therapeutics.

SUBMITTER: Yang S

PROVIDER: S-EPMC6060862 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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Prediction and characterization of a novel hemocyanin-derived antimicrobial peptide from shrimp Litopenaeus vannamei.

Yang Shen S Huang He H Wang Fan F Aweya Jude Juventus JJ Zheng Zhihong Z Zhang Yueling Y

Amino acids 20180504 8

Hemocyanin, the multifunctional glycoprotein in the hemolymph of invertebrates, can generate various antimicrobial peptides (AMPs). Given the rising interest in the use of natural therapeutic agents such as AMPs, alternative and more efficient methods for their generation are being explored. In this work, free online software was first applied to predict the generation of antimicrobial peptides from the large subunit of Litopenaeus vannamei hemocyanin. Twenty potential antimicrobial peptides ran ...[more]

PMID: 29728914

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Project description:The functional diversity of crustacean hemocyanins is broad, encompassing O2 delivery, innate immune response, metabolite storage, and osmolyte balance, all in a heterogeneous protein structure. As such, the sequence diversity of this class of proteins and its subunit composition are the focus of many studies on crustacean adaptation to environmental challenges. Recent transcriptomic and genomic sequencing on the Pacific whiteleg shrimp Litopenaeus vannamei has identified unique isoforms of hemocyanin including an ancestral β-type subunit thought to be lost in penaeid shrimp. However, it is unknown the degree to which these isoforms are translated as proteins, and whether they differ in function. The present study uses proteomic approaches to characterize the protein-level abundance and organization of these hemocyanin isoforms within their native oligomeric structures. Fractions of each hemocyanin oligomeric form were purified by size-exclusion high performance liquid chromatography for identification of subunit isoforms using tandem mass spectrometry at <1% protein false discovery rate. Relative abundances of hemocyanin oligomers and monomeric subunits from hemolymph and fractions were also quantified by polyacrylamide gel electrophoresis with and without denaturation for comparison of relative abundance. Ten hemocyanin isoforms were identified by tandem mass spectrometry in both hemocyanin oligomer fractions including a single small subunit, eight large subunits, and the first protein-level evidence of a β-type subunit in penaeid shrimp. Hemocyanin subunits were organized primarily as hexamers (95-99% relative abundance) as opposed to dodecamers. Hexamers utilized a significantly higher ratio (2.05:1) of small subunit to large subunit compared to dodecamers (1.04:1), and the relative abundances of the large subunit isoforms was dominated by HcL1 in both fractions. The ability to distinguish and quantify hemocyanin isoforms within oligomeric structures will aid future studies linking hemocyanin genes to function and physiology as well as offer insight into the evolutionary history of crustaceans.

Project description:It has long been viewed that invertebrates rely exclusively upon a wide variety of innate mechanisms for protection from disease and parasite invasion and lack any specific acquired immune mechanisms comparable to those of vertebrates. Recent findings, however, suggest certain invertebrates may be able to mount some form of specific immunity, termed 'specific immune priming', although the mechanism of this is not fully understood (see Textbox S1). In our initial experiments, either formalin-inactivated Vibrio harveyi or sterile saline were injected into the main body cavity (haemocoel) of juvenile shrimp (Litopenaeus vannamei). Haemocytes (blood cells) from V. harveyi-injected shrimp were collected 7 days later and incubated with a 1:1 mix of V. harveyi and an unrelated gram positive bacterium, Bacillus subtilis. Haemocytes from 'vaccinated' shrimp showed elevated levels of phagocytosis of V. harveyi, but not B. subtilis, compared with those from saline-injected (non-immunised) animals. The increased phagocytic activity was characterised by a significant increase in the percentage of phagocytic cells. When shrimp were injected with B. subtilis rather than vibrio, there was no significant increase in the phagocytic activity of haemocytes from these animals in comparison to the non-immunised (saline injected) controls. Whole haemolymph (blood) from either 'immunised' or non-immunised' shrimp was shown to display innate humoral antibacterial activity against V. harveyi that was absent against B. subtilis. However, there was no difference in the potency of antibacterial activity between V. harveyi-injected shrimp and control (saline injected) animals showing that 'vaccination' has no effect on this component of the shrimp's immune system. These results imply that the cellular immune system of shrimp, particularly phagocytosis, is capable of a degree of specificity and shows the phenomenon of 'immune priming' reported by other workers. However, in agreement with other studies, this phenomenon is not universal to all potential pathogens.

Dataset Information

Prediction and characterization of a novel hemocyanin-derived antimicrobial peptide from shrimp Litopenaeus vannamei.

Publications

Prediction and characterization of a novel hemocyanin-derived antimicrobial peptide from shrimp Litopenaeus vannamei.

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