Molecular Cloning, Expression, Purification, and Functional Characterization of Palustrin-2CE, an Antimicrobial Peptide of<i>Rana chensinensis</i>

Sun, Yan; Li, Qian; Li, Zhi; Yuan, Zhongwei; Jie, Zhao; Wang, Lin

doi:10.1271/bbb.110672

Cited by 16 publications

(6 citation statements)

References 30 publications

(35 reference statements)

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“…In this study, modified thioredoxin A was used as a fusion partner that promoted the correct disulfide bond formation and masked the toxic effects of AMPs. Previously, several frog AMPs bearing Rana-box were obtained in the heterologous Escherichia coli expression system [ 34 , 35 , 36 ]. A high proportion of a fusion protein in reference to total cell protein was achieved with the use of thioredoxin A or glutathione S-transferase (GST).…”

Section: Resultsmentioning

confidence: 99%

Novel Antimicrobial Peptides from the Arctic Polychaeta Nicomache minor Provide New Molecular Insight into Biological Role of the BRICHOS Domain

et al. 2018

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Endogenous antimicrobial peptides (AMPs) are among the earliest molecular factors in the evolution of animal innate immunity. In this study, novel AMPs named nicomicins were identified in the small marine polychaeta Nicomache minor in the Maldanidae family. Full-length mRNA sequences encoded 239-residue prepropeptides consisting of a putative signal sequence region, the BRICHOS domain within an acidic proregion, and 33-residue mature cationic peptides. Nicomicin-1 was expressed in the bacterial system, and its spatial structure was analyzed by circular dichroism and nuclear magnetic resonance spectroscopy. Nicomicins are unique among polychaeta AMPs scaffolds, combining an amphipathic N-terminal α-helix and C-terminal extended part with a six-residue loop stabilized by a disulfide bridge. This structural arrangement resembles the Rana-box motif observed in the α-helical host-defense peptides isolated from frog skin. Nicomicin-1 exhibited strong in vitro antimicrobial activity against Gram-positive bacteria at submicromolar concentrations. The main mechanism of nicomicin-1 action is based on membrane damage but not on the inhibition of bacterial translation. The peptide possessed cytotoxicity against cancer and normal adherent cells as well as toward human erythrocytes.

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Section: Resultsmentioning

confidence: 99%

Novel Antimicrobial Peptides from the Arctic Polychaeta Nicomache minor Provide New Molecular Insight into Biological Role of the BRICHOS Domain

et al. 2018

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“…The E. coli recombinant expression system is a suitable choice for large-scale production, due to its easy culture, fast growth and effective prevention of bacterial contamination (25). Numerous AMPs have been prepared successfully in E. coli, including brevinin-2R (26) and palusterin-2CE (27). A number of fusion partners have been used to express AMPs, including maltose-binding protein, thioredoxin and green fluorescent protein (28–30), to avoid toxicity and proteolysis of AMPs and increase their expression levels in E. coli (17,24).…”

Section: Discussionmentioning

confidence: 99%

Expression and characterization of cecropinXJ, a bioactive antimicrobial peptide from Bombyx mori (Bombycidae, Lepidoptera) in Escherichia coli

Zhang

Liu

et al. 2013

Experimental and Therapeutic Medicine

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Insect antimicrobial peptides (AMPs) have a broad antimicrobial spectrum. To aid the characterization of the gene function and further applications, we cloned the gene of cecropinXJ into the prokaryotic expression vector pET32a and expressed cecropinXJ in Escherichia coli BL2l (DE3). Following induction by isopropyl-β-D-thiogalactoside (IPTG), a 25 kDa fusion peptide of cecropinXJ with a tagged thioredoxin (Trx) protein was highly expressed in E. coli. The yield was 10 mg/l culture medium following purification on nickel-nitrilotriacetic acid (Ni-NTA) metal affinity chromatography matrices. The purified recombinant antibacterial peptide, cecropinXJ, retained a high stability against Staphylococcus aureus over a temperature range from 4 to 100°C and a pH range from pH 2.0 to 12.0. The minimum inhibitory concentration (MIC) of the fusion protein against S. aureus was 0.4 μM. The recombinant cecropinXJ is also cytotoxic to several types of human cancer cells.

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“…In addition to the easy and fast growth of E. coli cultures, most of the antimicrobial peptides bear no posttranslational modifications which allow for the use of E. coli as a suitable choice for large-scale production (Huang et al 2009). Several AMPs have already been successfully expressed in E. coli such as brevinin-2R (Mehrnejad et al 2008) and palusterin-2CE (Sun et al 2012). A number of fusion partners have been used to express AMPs including maltose-binding protein, thioredoxin, and green fluorescent protein (Kapust and Waugh 1999;Lee et al 1998;Skosyrev et al 2003).…”

Section: Discussionmentioning

confidence: 99%

Molecular cloning and expression of ranalexin, a bioactive antimicrobial peptide from Rana catesbeiana in Escherichia coli and assessments of its biological activities

Aleinein

Hamoud

Schäfer

2012

Appl Microbiol Biotechnol

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The coding sequence, which corresponds to the mature antimicrobial peptide ranalexin from the frog Rana catesbeiana, was chemically synthesized with preferred codons for expression in Escherichia coli. It was cloned into the vector pET32c (+) to express a thioredoxin-ranalexin fusion protein which was produced in soluble form in E. coli BL21 (DE3) induced under optimized conditions. After two purification steps through affinity chromatography, about 1 mg of the recombinant ranalexin was obtained from 1 L of culture. Mass spectrometrical analysis of the purified recombinant ranalexin demonstrated its identity with ranalexin. The purified recombinant ranalexin is biologically active. It showed antibacterial activities similar to those of the native peptide against Staphylococcus aureus, Streptococcus pyogenes, E. coli, and multidrug-resistant strains of S. aureus with minimum inhibitory concentration values between 8 and 128 μg/ml. The recombinant ranalexin is also cytotoxic in HeLa and COS7 human cancer cells (IC50 = 13-15 μg/ml).

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Molecular Cloning, Expression, Purification, and Functional Characterization of Palustrin-2CE, an Antimicrobial Peptide ofRana chensinensis

Cited by 16 publications

References 30 publications

Novel Antimicrobial Peptides from the Arctic Polychaeta Nicomache minor Provide New Molecular Insight into Biological Role of the BRICHOS Domain

Novel Antimicrobial Peptides from the Arctic Polychaeta Nicomache minor Provide New Molecular Insight into Biological Role of the BRICHOS Domain

Expression and characterization of cecropinXJ, a bioactive antimicrobial peptide from Bombyx mori (Bombycidae, Lepidoptera) in Escherichia coli

Molecular cloning and expression of ranalexin, a bioactive antimicrobial peptide from Rana catesbeiana in Escherichia coli and assessments of its biological activities

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