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Last updated: 13th August 2010

A defensin antimicrobial peptide from the venoms of Nasonia vitripennis.
Ye J, Zhao H, Wang H, Bian J, Zheng R
Intensive Care Unit of Taizhou People´s Hospital, Taizhou, Jiangsu 225300, China.
Toxicon 2010 Aug;56 (1):101-6
Although many antimicrobial components (i.e. antimicrobial peptides) have been found in many social Hymenoptera venoms, no antimicrobial compound is purified and characterized from parasitic Hymenoptera. From the venoms of the ectoparasitic wasp, Nasonia vitripennis, a defensin-like antimicrobial peptide named defensin-NV was purified and characterized. Defensin-NV is composed of 52 amino acid residues including 6 cysteines forming 3 disulfide bridges. Its amino acid sequence is VTCELLMFGGVVGDSACAANCLSMGKAGGSCNGGLCDCRKTTFKELWDKRFG. By BLAST search, defensin-NV showed significant sequence similarity to other insect defensin antimicrobial peptides. Defensin-NV exerted strong antimicrobial activity against tested microorganisms including Gram-positive bacteria, Gram-negative bacteria and fungi. The cDNA encoding defensin-NV was cloned from the venom reservoir cDNA library of N. vitripennis. The current work firstly purified and characterized an antimicrobial peptide from parasitic Hymenoptera.
Evolution of a cluster of innate immune genes (beta-defensins) along the ancestral lines of chicken and zebra finch.
Hellgren O, Ekblom R
Edward Grey Institute, Department of Zoology, South Parks Road, Oxford, OX1 3PS, UK.
Immunome Res 2010 ;6 3
BACKGROUND: Avian beta-defensins (AvBDs) represent a group of innate immune genes with broad antimicrobial activity. Within the chicken genome, previous work identified 14 AvBDs in a cluster on chromosome three. The release of a second bird genome, the zebra finch, allows us to study the comparative evolutionary history of these gene clusters between from two species that shared a common ancestor about 100 million years ago. RESULTS: A phylogenetic analysis of the beta-defensin gene clusters in the chicken and the zebra finch identified several cases of gene duplication and gene loss along their ancestral lines. In the zebra finch genome a cluster of 22 AvBD genes were identified, all located within 125 Kbp on chromosome three. Ten of the 22 genes were found to be highly conserved with orthologous genes in the chicken genome. The remaining 12 genes were all located within a cluster of 58 Kbp and are suggested to be a result of recent gene duplication events that occurred after the galliformes- passeriformes split (G-P split). Within the chicken genome, AvBD6 was found to be a duplication of AvBD7, whereas the gene AvDB14 seems to have been lost along the ancestral line of the zebra finch. The duplicated beta-defensin genes have had a significantly higher accumulation of non-synonymous over synonymous substitutions compared to the genes that have not undergone duplication since the G-P split. The expression patterns of avian beta-defensin genes seem to be well conserved between chicken and zebra finch. CONCLUSION: The genomic comparisons of the beta-defensins gene clusters of the chicken and zebra finch illuminate the evolutionary history of this gene complex. Along their ancestral lines, several gene duplication events have occurred in the passerine line after the galliformes-passeriformes split giving rise to 12 novel genes compared to a single duplication event in the galliformes line. After the duplication events, the duplicated genes have been subject to a relaxed selection pressure compared to the non-duplicated genes, thus supporting models of evolution by gene duplication.
Expression of defensins in non-infected araneomorph spiders.
Baumann T, Kuhn-Nentwig L, Largiadèr CR, Nentwig W
Institute of Ecology and Evolution, University of Bern, Baltzerstrasse 6, 3012, Bern, Switzerland.
Cell Mol Life Sci. 2010 Aug;67(15):2643-51.
Defensins are a major family of antimicrobial peptides found throughout the phylogenetic tree. From the spider species: Cupiennius salei, Phoneutria reidyi, Polybetes pythagoricus, Tegenaria atrica, and Meta menardi, defensins belonging to the 'ancestral' class of invertebrate defensins were cloned and sequenced. The deduced amino acid sequences contain the characteristic six cysteines of this class of defensins and reveal precursors of 60 or 61 amino acid residues. The mature peptides consist of 37 amino acid residues, showing up to 70% identities with tick and scorpion defensins. In C. salei, defensin mRNA was found to be constitutively expressed in hemocytes, ovaries, subesophageal nerve mass, hepatopancreas, and muscle tissue. This is the first report presenting and comparing antimicrobial peptides belonging to the family of defensins from spiders.
Validation of a quantitative assay for human neutrophil peptide-1, -2, and -3 in human plasma and serum by liquid chromatography coupled to tandem mass spectrometry.
van den Broek I, Sparidans RW, Schellens JH, Beijnen JH
Utrecht University, Faculty of Science, Department of Pharmaceutical Sciences, Section of Biomedical Analysis, Division of Drug Toxicology, Sorbonnelaan 16, 3584 CA Utrecht, The Netherlands.
J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 2010 May;878 (15-16):1085-92
A quantitative assay for simultaneous measurement of individual human neutrophil peptide-1, -2 and -3 concentrations will aid in exploring the potential of these antimicrobial peptides as biomarkers for various diseases. Therefore, a liquid chromatography-tandem mass spectrometry method has been developed and validated to allow separate quantification of the three human neutrophil peptides in human plasma and serum. Plasma and serum samples (100 microl) were deproteinized by precipitation, followed by chromatographic separation on a Symmetry 300 C18 column (50 mm x 2.1mm I.D., particle size 3.5 microm), using a water-methanol gradient containing 0.25% (v/v) formic acid and human alpha-defensin 5 as internal standard. Tandem mass spectrometric detection was performed on a triple quadrupole mass spectrometer equipped with electrospray ionization. Despite low fragmentation efficiency of the antimicrobial peptides, multiple reaction monitoring was used for detection, though selecting the quaternary charged ions as both precursor and product. The method was linear for concentrations between 5 and 1000 ng/ml with a limit of detection around 3 ng/ml for all peptides. Intra- and inter-assay precisions were 14.8 and 19.1%, respectively, at the lowest measured endogenous concentration (6.4 ng/ml of HNP-1 in plasma), representing the lower limit of quantification of the assay. Recoveries of HNP-1, -2 and -3 from plasma and serum ranged between 85 and 128%. Analysis of serum samples from intensive care patients showed average concentrations of 362, 570 and 143 ng/ml for HNP-1, -2 and -3, respectively.
Proteomic analysis of common bean seed with storage protein deficiency reveals up-regulation of sulfur-rich proteins and starch and raffinose metabolic enzymes, and down-regulation of the secretory pathway.
Marsolais F, Pajak A, Yin F, Taylor M, Gabriel M, Merino DM, Ma V, Kameka A, Vijayan P, Pham H, Huang S, Rivoal J, Bett K, Hernández-Sebastià C, Liu Q, Bertrand A, Chapman R
Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, London, ON, Canada.
J Proteomics. 2010 Jun 16;73(8):1587-600.
A deficiency in major seed storage proteins is associated with a nearly two-fold increase in sulfur amino acid content in genetically related lines of common bean (Phaseolus vulgaris). Their mature seed proteome was compared by an approach combining label-free quantification by spectral counting, 2-DE, and analysis of selective extracts. Lack of phaseolin, phytohemagglutinin and arcelin was mainly compensated by increases in legumin, alpha-amylase inhibitors and mannose lectin FRIL. Along with legumin, albumin-2, defensin and albumin-1 were major contributors to the elevated sulfur amino acid content. Coordinate induction of granule-bound starch synthase I, starch synthase II-2 and starch branching enzyme were associated with minor alteration of starch composition, whereas increased levels of UDP-glucose 4-epimerase were correlated with a 30% increase in raffinose content. Induction of cell division cycle protein 48 and ubiquitin suggested enhanced ER-associated degradation. This was not associated with a classical unfolded protein response as the levels of ER HSC70-cognate binding protein were actually reduced in the mutant. Repression of rab1 GTPase was consistent with decreased traffic through the secretory pathway. Collectively, these results have implications for the nutritional quality of common bean, and provide information on the pleiotropic phenotype associated with storage protein deficiency in a dicotyledonous seed.
Molecular analysis and recombinant expression of bovine neutrophil beta-defensin 12 and its antimicrobial activity.
Wu J, Wang C, He H, Hu G, Yang H, Gao Y, Zhong J
Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, No. 159-1, Industry North Road, 250100, Jinan, Shandong Province, People´s Republic of China.
Molecular biology reports 2010 Mar
To analyze molecular characteristics and antimicrobial activity of bovine neutrophil beta-defensin 12 (BNBD12), Pro-BNBD12 gene was amplified from Chinese Holstein cow using reverse transcription polymerase chain reaction. Based on the sequence of mature BNBD12 and codon preference in E. coli, a modified mature BNBD12 cDNA was synthesized, cloned into pET32a (+) vector and expressed in E. coli BL21 as a 26 kD fusion protein after isopropyl beta-D: -1-thiogalactopyranoside induction. The expressed mature BNBD12 accounted for 21.4% of total protein. 0.05 mg/ml purified BNBD12 had antimicrobial activity against both E. coli and S. aureus in vitro assayed by agar diffusion method. Electron microscopy found that BNBD12 treatment of E. coli and S. aureus could induce cell content leakage. Taking together, BNBD12 protein was successfully expressed in E. coli and it has antimicrobial activity against both Gram-positive and negative bacteria and potentials in control of mastitis clinically.
Expression of antimicrobial peptides in cutaneous infections after skin surgery.
Kesting MR, Stoeckelhuber M, Hölzle F, Mücke T, Neumann K, Woermann K, Jacobsen F, Steinstraesser L, Wolff KD, Loeffelbein DJ, Rohleder NH
Department of Oral and Maxillofacial Surgery, Technische Universität München, Ismaninger Str. 22, D-81675 Munich, Germany.
Br J Dermatol. 2010 Jul;163(1):121-7.
BACKGROUND: Increasing numbers of antibiotics have lost efficiency because of bacterial resistance. The consequences can be severe when surgical wounds become infected during postoperative care. Natural peptide antibiotics, the so-called host defence peptides (HDPs), have been investigated since the 1990s in a search for alternative treatment strategies. HDPs build up a protection shield against pathological microorganisms, especially in human epithelium. The use of HDPs is currently being discussed as a new antimicrobial therapeutic strategy. Accordingly, a profound knowledge of the quantitative relationships of the effectors is essential. OBJECTIVES: To evaluate differences in HDP expression between postoperatively inflamed and healthy epithelium. METHODS: Expression profiles of the genes encoding HDP human beta-defensin (hBD)-1 (DEFB1, previously known as HBD-1), hBD-2 (DEFB4A, previously known as HBD-2), hBD-3 (DEFB103A, previously known as HBD-3) and psoriasin (S100A7) were assessed in samples of surgical wound healing disorders (n = 27) and healthy epithelium (n = 16) by using real-time polymerase chain reaction. Immunohistochemical staining was performed in the same samples. RESULTS: A significant overexpression of DEFB4A (P < 0.001), DEFB103A (P = 0.001) and S100A7 (P < 0.001) was found in cutaneous surgical site infections. Immunohistochemistry revealed intensely elevated protein levels of psoriasin in infected wounds, and differences in distribution with respect to the epithelial layers. CONCLUSIONS: The study demonstrates upregulated mRNA expression and protein levels of HDPs in postoperatively inflamed epithelium. The results may be a starting point for novel pharmacological treatments.
Skin bacteria after chlorhexidine exposure-is there a difference in response to human beta-Defensin-3?
Reichel M, Heisig A, Heisig P, Kampf G
BODE Chemie GmbH, Scientific Affairs, Melanchthonstr. 27, 22525, Hamburg, Germany.
Eur J Clin Microbiol Infect Dis. 2010 Jun;29(6):623-32.
We investigated whether exposure to sub-lethal concentrations of chlorhexidine digluconate (CHG) changed the response of five Staphylococcus spp. to human beta-Defensin-3 (hBD-3). The change in response for each strain was determined in vitro with time-kill experiments in suspension by comparing the mean log(10) reduction caused by hBD-3 at 1.5 and 3 h in exposed and non-exposed bacteria. The identity of staphylococcal species was verified by DNA sequence homology in the gyrA genes in comparison with reference strains. Baseline sub-lethal concentrations allowing visible bacterial growth were between 0.0625 and 0.25 mug/ml. Sub-lethal CHG concentrations increased within 3 days in two isolates. For S. capitis 19/2, CHG-exposed cells were less susceptible to 0.5 mug/ml hBD-3 (log(10) reduction 0.78 versus 2.06 at 1.5 h; p < 0.001; t-test). For S. aureus, however, CHG-exposed cells were more susceptible to 1 mug/ml hBD-3. The observed changes between CHG-exposed and non-exposed cells did not indicate a general trend in response to hBD-3. Overall, we found no consistent evidence that 3 days of exposure to CHG changed the response of five Staphylococcus spp. to hBD-3. The use of CHG for skin antisepsis is, based on our data, unlikely to change the natural defence activity of hBD-3.
Antifungal activity of recombinant mouse beta-defensin 3.
Jiang Y, Wang Y, Wang B, Yang D, Yu K, Yang X, Liu F, Jiang Z, Li M
Department of Microbiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Lett Appl Microbiol. 2010 May;50(5):468-73.
Abstract Aims: To identify the presence of mouse beta-defensin 3 (Mbd3) (the human homologue of beta-defensin 2) in different tissues and to define the antimicrobial properties of recombinant MBD3 (rMBD3) against a panel of human pathogens. Methods and Results: Mbd3 gene expression in different mouse tissues before or after lipopolysaccharide (LPS) injection was compared by semi-quantitative RT-PCR. This analysis demonstrated that epithelial and mucosal tissues expressed Mbd3 independent of LPS stimulation. Evaluation of the antimicrobial properties of recombinant rMBD3 was determined by assessing the median inhibition concentration (IC(50)), minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC)/minimal fungicidal concentration (MFC) against various human pathogens. Conclusion: Mbd3 gene expression by epithelial and mucosal tissues suggested that MBD3 likely plays an early defensive role against microbial infections. This activity was most significant against filamentous fungi. Significance and Impact of the Study: The data presented in this report suggested that formulations containing rMBD3 and related molecules could serve to treat fungal and bacterial infections.
Human beta-defensin-3 up-regulates cyclooxygenase-2 expression and prostaglandin E synthesis in human gingival fibroblasts.
Chotjumlong P, Khongkhunthian S, Ongchai S, Reutrakul V, Krisanaprakornkit S
Department of Oral Biology and Diagnostic Sciences and Center of Excellence for Innovation in Chemistry, Chiang Mai University, Chiang Mai, Thailand.
J Periodontal Res. 2010 Aug;45(4):464-70.
BACKGROUND AND OBJECTIVE: Oral epithelial cells express three antimicrobial peptide human beta-defensins (hBDs) that have previously been demonstrated to exert proinflammatory effects on various immune cells. We wanted to examine whether hBDs could induce cyclooxygenase-2 (COX-2) expression and prostaglandin E(2) (PGE(2)) synthesis in non-immune cells, such as human gingival fibroblasts. MATERIAL AND METHODS: Cultured fibroblasts were treated with different concentrations of hBD-1, -2, -3 or interleukin-1 beta, as a positive control, for various times, in the presence or absence of NS-398, a specific COX-2 inhibitor. The levels of COX-1 and COX-2 mRNA expression were analyzed using RT-PCR and real-time PCR. Whole cell lysates were analyzed for COX-1 and COX-2 protein expression by western blotting. Cell-free culture supernatants were assayed for PGE(2) levels by ELISA. The lactate dehydrogenase assay was performed to determine the cytotoxicity of hBDs. RESULTS: Ten and 40 microg/mL of hBD-3 up-regulated COX-2 mRNA and protein expression, consistent with COX-2 up-regulation by interleukin-1 beta, whereas hBD-1 and hBD-2 did not. However, COX-1 mRNA and protein were constitutively expressed. The time-course study revealed that hBD-3 up-regulated COX-2 mRNA and protein expression at 6 and 12 h, respectively. Consistent with COX-2 up-regulation, 10 and 40 microg/mL of hBD-3 significantly increased PGE(2) levels in cell-free culture supernatants (p < 0.05), and this was inhibited by NS-398 in a dose-dependent manner. Neither of the hBD concentrations tested in this study was toxic to the cells. CONCLUSION: These findings indicate that epithelial human beta-defensin-3 functions as a proinflammatory mediator in controlling arachidonic acid metabolism in underlying fibroblasts.