scholarly journals Expression and functional characterization of a novel antimicrobial peptide: human beta-defensin118

2020 ◽  
Author(s):  
Qian Lin ◽  
KunHong Xie ◽  
Daiwen Chen ◽  
Bing Yu ◽  
Xiangbing Mao ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Functional Characterization ◽  
Antimicrobial Activities ◽  
Bacterial Strains ◽  
E Coli ◽  
Gram Positive Bacteria ◽  
Sds Page ◽  
Novel Host ◽  
Late Apoptosis

Abstract Background: β-defensin 118 (DEFB118 ) is a novel host defence peptide (HDP) identified in human. To evaluate its potentials for future utilization, the DEFB118 gene was expressed in Escherichia coli ( E. coli ) and the recombinant protein was fully characterized. Methods: The DEFB118 protein was obtained by heterologous expression using E. coli Rosetta (DE3). Antibacterical activity of DEFB118 were determined by using various bacterial strains. IPEC-J cells challenged by E. coli K88 were used to determine its influences on inflammatory responses. Results: The E. coli transformants yielded more than 250 mg/mL D EFB118 protein after 4 h induction by 1.0 mM IPTG. The DEFB118 was estimated by SDS-PAGE to be 30 kDa, and MALDI-TOF analysis verified it is a human β-defensin 118. Importantly, the DEFB118 showed antimicrobial activities against both Gram-negative bacteria ( E. coli K88 and E. coli DH5α) and Gram-positive bacteria ( S. aureus and B. subtilis ), with a minimum inhibitory concentration (MIC) of 4 μg/mL. Hemolytic assays showed that DEFB118 had no detrimental impact on cell viability. Additionally, DEFB118 was found to elevate the viability of IPEC-J2 cells upon E. coli K88 challenge. Moreover, DEFB118 significantly decreased cell apoptosis in the late apoptosis phase and down-regulated the expression of inflammatory cytokines such as the IL-1β and TNF-a in the IPEC-J2 cells exposure to E. coli K88. Conclusions: These results suggested a novel function of the mammalian defensins, and the anti-bacterial and anti-inflammatory properties of DEFB118 may allow it a potential substitute for conventionally used antibiotics or drugs.

scholarly journals Expression and Functional Characterization of a Novel Antimicrobial Peptide: Human Beta-Defensin 118

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
Qian Lin ◽  
Kunhong Xie ◽  
Daiwen Chen ◽  
Bing Yu ◽  
Xiangbing Mao ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Functional Characterization ◽  
Antimicrobial Activities ◽  
Bacterial Strains ◽  
E Coli ◽  
Gram Positive Bacteria ◽  
Tnf Α ◽  
Novel Host ◽  
Host Defense Peptide ◽  
Late Apoptosis

Purpose. β-Defensin 118 (DEFB118) is a novel host defense peptide (HDP) identified in humans. To evaluate its potentials for future utilization, the DEFB118 gene was expressed in Escherichia coli (E. coli) and the recombinant protein was fully characterized. Methods. The DEFB118 protein was obtained by heterologous expression using E. coli Rosetta (DE3). Antibacterial activity of DEFB118 was determined by using various bacterial strains. IPEC-J cells challenged by E. coli K88 were used to determine its influences on inflammatory responses. Results. The E. coli transformants yielded more than 250 μg/mL DEFB118 protein after 4 h induction by 1.0 mM IPTG. The DEFB118 was estimated by SDS-PAGE to be 30 kDa, and MALDI-TOF analysis verified that it is a human β-defensin 118. Importantly, the DEFB118 showed antimicrobial activities against both Gram-negative bacteria (E. coli K88 and E. coli DH5α) and Gram-positive bacteria (S. aureus and B. subtilis), with a minimum inhibitory concentration (MIC) of 4 μg/mL. Hemolytic assays showed that DEFB118 had no detrimental impact on cell viability. Additionally, DEFB118 was found to elevate the viability of IPEC-J2 cells upon E. coli K88 challenge. Moreover, DEFB118 significantly decreased cell apoptosis in the late apoptosis phase and downregulated the expression of inflammatory cytokines such as IL-1β and TNF-α in IPEC-J2 cell exposure to E. coli K88. Conclusions. These results suggested a novel function of the mammalian defensins, and the antibacterial and anti-inflammatory properties of DEFB118 may allow it as a potential substitute for conventionally used antibiotics or drugs.

scholarly journals Functional Characterization of Porcine NK-Lysin: A Novel Immunomodulator That Regulates Intestinal Inflammatory Response

Molecules ◽  
2021 ◽  
Vol 26 (14) ◽  
pp. 4242
Author(s):  
Qian Lin ◽  
Qingqing Fu ◽  
Daiwen Chen ◽  
Bing Yu ◽  
Yuheng Luo ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Functional Characterization ◽  
Antibacterial Activities ◽  
Bacterial Strains ◽  
E Coli ◽  
Gram Positive Bacteria ◽  
Tnf Α ◽  
Sds Page ◽  
Junction Protein

Porcine NK-Lysine (PNKL) is a new antimicrobial peptide (AMP) identified in the small intestine. In this study, PNKL protein was obtained through heterologous expression in Escherichia coli and was estimated by SDS-PAGE at 33 kDa. The antibacterial activities of PNKL were determined using various bacterial strains and showed broad-spectrum antimicrobial activity against Gram-negative and Gram-positive bacteria. Furthermore, E. coli K88-challenged IPEC-J2 cells were used to determine PNKL influences on inflammatory responses. Hemolytic assays showed that PNKL had no detrimental impact on cell viability. Interestingly, PNKL elevated the viability of IPEC-J2 cells exposure to E. coli K88. PNKL significantly decreased the cell apoptosis rate, and improved the distribution and abundance of tight junction protein ZO-1 in IPEC-J2 cells upon E. coli K88-challenge. Importantly, PNKL not only down regulated the expressions of inflammatory cytokines such as the IL-6 and TNF-α, but also down regulated the expressions of NF-κB, Caspase3, and Caspase9 in the E. coli K88-challenged cells. These results suggest a novel function of natural killer (NK)-lysin, and the anti-bacterial and anti-inflammatory properties of PNKL may allow it a potential substitute for conventionally used antibiotics or drugs.

scholarly journals Fabrication of Gelatin/Chitosan/Zeolite Composite Packaging Film for Enhancing the Quality of Fresh Meat During Storage

2020 ◽  
Vol 2 (1) ◽  
pp. 53
Keyword(s):  
Food Packaging ◽  
Functional Characterization ◽  
Antimicrobial Activities ◽  
Meat Processing ◽  
Physico Chemical ◽  
Sds Page ◽  
Zeolite Composite ◽  
Biocomposite Film

Valorisation of fishery by-products for the extraction of gelatin-like biopolymers is an active area of research presently due to their significant food and biomedical applications. In the present study, gelatin was extracted from the discarded portions of the Amphioctopusmarginatus. One of the major octopus species landed near the Kerala coast. Physico-chemical and functional characterization of the extracted gelatin was done based on SDS- PAGE, UV-Vis spectrophotometer, FTIR, XRD, HPLC, gelling and emulsifying capacities. Later, biocomposite film based on the gelatin/chitosan/zeolite was developed and characterized according to its solubility, opacity, spectroscopic analysis, antioxidative and antimicrobial activities. The results showed the dimensions of improving the properties of biopolymer films in terms of its ability to prevent the oxidative lipid changes, protein degradation and microbial spoilage in packaged fresh chicken meat over 9 days of refrigerated storage. The gelatin/chitosan/zeolite composite film thus developed has the potential to serve as a promising bio-based packaging material in the food packaging and meat processing industries.

scholarly journals Molecular Cloning, Expression, Purification, and Functional Characterization of Dammarenediol Synthase fromPanax ginseng

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Wei Hu ◽  
Ning Liu ◽  
Yuhua Tian ◽  
Lianxue Zhang
Keyword(s):  
Cdna Library ◽  
Functional Characterization ◽  
Vital Role ◽  
Ginseng Root ◽  
Phage Library ◽  
E Coli ◽  
Sds Page ◽  
Ginsenoside Content ◽  
Dammarenediol Synthase

The objective of this study is to clone and charecterize the expression of dammarenediol synthase gene and then to determine the relationship between the expression of dammarenediol synthase gene that is involved in the ginsenoside biosynthetic pathway and the ginsenoside content. A cDNA phage library was constructed from a five-year-old ginseng root. The cDNA library was screened for the dammarenediol synthase gene by using its specific primers. It was further cloned and expressed in pET-30a vector. The recombinant plasmid pET-30a-DS was expressed in RosettaE. coli. The recombinant DS protein was purified by affinity chromatography. The production of dammarenediol was detected by liquid chromatography-mass spectrometry (LC-MS). Results showed that dammarenediol synthase gene was cloned from the cDNA library and was expressed in RosettaE. coliand the SDS-PAGE analysis showed the presence of purified DS protein. LS-MS showed the activity of DS protein, as the protein content increases the dammarenediol increases. Our results indicate that the recombinant dammarenediol synthase protein could increase the production of dammarenediol and the expression of DS played a vital role in the biosynthesis of ginsenosides inP. ginseng.

scholarly journals Functional identification of ygiP as a positive regulator of the ttdA-ttdB-ygjE operon

Microbiology ◽  
2006 ◽  
Vol 152 (7) ◽  
pp. 2129-2135 ◽  
Author(s):  
Taku Oshima ◽  
Francis Biville
Keyword(s):  
Functional Characterization ◽  
Anaerobic Growth ◽  
Functional Identification ◽  
New Members ◽  
E Coli ◽  
Inner Membrane Protein ◽  
Tartrate Dehydratase

Functional characterization of unknown genes is currently a major task in biology. The search for gene function involves a combination of various in silico, in vitro and in vivo approaches. Available knowledge from the study of more than 21 LysR-type regulators in Escherichia coli has facilitated the classification of new members of the family. From sequence similarities and its location on the E. coli chromosome, it is suggested that ygiP encodes a lysR regulator controlling the expression of a neighbouring operon; this operon encodes the two subunits of tartrate dehydratase (TtdA, TtdB) and YgiE, an integral inner-membrane protein possibly involved in tartrate uptake. Expression of tartrate dehydratase, which converts tartrate to oxaloacetate, is required for anaerobic growth on glycerol as carbon source in the presence of tartrate. Here, it has been demonstrated that disruption of ygiP, ttdA or ygjE abolishes tartrate-dependent anaerobic growth on glycerol. It has also been shown that tartrate-dependent induction of the ttdA-ttdB-ygjE operon requires a functional YgiP.

scholarly journals Conjuring up a ghost: structural and functional characterization of FhuF, a ferric siderophore reductase from E. coli

2021 ◽  
Author(s):  
I. B. Trindade ◽  
G. Hernandez ◽  
E. Lebègue ◽  
F. Barrière ◽  
T. Cordeiro ◽  
...  
Keyword(s):  
Functional Characterization ◽  
Paramagnetic Nmr ◽  
E Coli ◽  
The Past ◽  
Internal Cavities ◽  
Structural Differences ◽  
Bond Strengths ◽  
Redox Bohr Effect ◽  
Micromolar Range

AbstractIron is a fundamental element for virtually all forms of life. Despite its abundance, its bioavailability is limited, and thus, microbes developed siderophores, small molecules, which are synthesized inside the cell and then released outside for iron scavenging. Once inside the cell, iron removal does not occur spontaneously, instead this process is mediated by siderophore-interacting proteins (SIP) and/or by ferric-siderophore reductases (FSR). In the past two decades, representatives of the SIP subfamily have been structurally and biochemically characterized; however, the same was not achieved for the FSR subfamily. Here, we initiate the structural and functional characterization of FhuF, the first and only FSR ever isolated. FhuF is a globular monomeric protein mainly composed by α-helices sheltering internal cavities in a fold resembling the “palm” domain found in siderophore biosynthetic enzymes. Paramagnetic NMR spectroscopy revealed that the core of the cluster has electronic properties in line with those of previously characterized 2Fe–2S ferredoxins and differences appear to be confined to the coordination of Fe(III) in the reduced protein. In particular, the two cysteines coordinating this iron appear to have substantially different bond strengths. In similarity with the proteins from the SIP subfamily, FhuF binds both the iron-loaded and the apo forms of ferrichrome in the micromolar range and cyclic voltammetry reveals the presence of redox-Bohr effect, which broadens the range of ferric-siderophore substrates that can be thermodynamically accessible for reduction. This study suggests that despite the structural differences between FSR and SIP proteins, mechanistic similarities exist between the two classes of proteins. Graphic abstract

Sa1806 Molecular and Functional Characterization of Fimh in Crohn's Disease Associated E. Coli

2013 ◽  
Vol 144 (5) ◽  
pp. S-310
Author(s):  
Brendan Chandler ◽  
Belgin Dogan ◽  
Ellen J. Scherl ◽  
Kenneth W. Simpson
Keyword(s):  
Crohn’S Disease ◽  
Crohn's Disease ◽  
Functional Characterization ◽  
E Coli

scholarly journals DPPH-Scavenging and Antimicrobial Activities of Asteraceae Medicinal Plants on Uropathogenic Bacteria

2020 ◽  
Vol 2020 ◽  
pp. 1-9 ◽  
Author(s):  
Phan-Canh Trinh ◽  
Le-Thi-Thanh Thao ◽  
Hoang-Tran-Viet Ha ◽  
TuAnh Nguyen
Keyword(s):  
Antimicrobial Agents ◽  
Natural Antioxidants ◽  
Ethyl Acetate Fraction ◽  
Antimicrobial Activities ◽  
Chrysanthemum Morifolium ◽  
Bacterial Strains ◽  
Ageratum Conyzoides ◽  
Potent Effect ◽  
E Coli ◽  
Dpph Scavenging

Asteraceae species were widely applied in traditional medicines in Asian countries as sources of natural antioxidants and antimicrobial agents. This study aimed to evaluate DPPH-scavenging capacities and antimicrobial activities of nine Asteraceae species collected from Southern Vietnam. Antioxidant and antimicrobial activities were determined by standard protocols. Essential oils from Ageratum conyzoides, Helianthus annuus, and Artemisia vulgaris indicated significant inhibitory effects on Staphylococcus aureus and Candida spp. Crude extracts and fractions from Taraxacum officinale, Chrysanthemum morifolium, A. conyzoides, and Tagetes erecta showed inhibitory ability on at least one testing bacterial strains including S. aureus, Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa. In a study on clinical isolates, ethyl acetate fraction from A. conyzoides flower displayed the most potent effect on uropathogenic E. coli and K. pneumoniae with MIC at 1.25–10 mg/ml and 5–12.5 mg/ml, respectively. DPPH-scavenging assay indicated that T. erecta extract had the lowest IC50 (17.280 μg/ml) and is 2.4 times higher than vitamin C (7.321 μg/ml). This study revealed that A. conyzoides has good potential against uropathogenic E. coli and K. pneumoniae, and therefore could be applied for prophylactic treatment of urinary infection.

scholarly journals Expression, secretion and functional characterization of three laccases in E. coli

2022 ◽  
Vol 7 (1) ◽  
pp. 474-480
Author(s):  
Yating Mo ◽  
Hou Ip Lao ◽  
Sau Wa Au ◽  
Ieng Chon Li ◽  
Jeremy Hu ◽  
...  
Keyword(s):  
Functional Characterization ◽  
E Coli

scholarly journals Viability of lactic acid bacteria in different components of Ogi with anti diarrhoeagenic E. coli activities

2019 ◽  
Vol 3 (6) ◽  
pp. 206-213
Author(s):  
Roseline Eleojo Kwasi ◽  
Iyanuoluwa Gladys Aremu ◽  
Qudus Olamide Dosunmu ◽  
Funmilola A. Ayeni
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Antimicrobial Properties ◽  
Culture Method ◽  
Antimicrobial Activities ◽  
Drying Methods ◽  
Bacterial Strains ◽  
E Coli ◽  
Log Reduction ◽  
Freeze Dried

Background: Ogi constitutes a rich source of lactic acid bacteria (LAB) with associated health benefits to humans through antimicrobial activities. However, the high viability of LAB in Ogi and its supernatant (Omidun) is essential. Aims: This study was carried out to assess the viability of LAB in various forms of modified and natural Ogi and the antimicrobial properties of Omidun against diarrhoeagenic E coli. Methods and Material: The viability of LAB was assessed in fermented Ogi slurry and Omidun for one month and also freeze-dried Ogi with and without added bacterial strains for two months. A further 10 days viability study of modified Omidun, refrigerated Omidun, and normal Ogi was performed. The antimicrobial effects of modified Omidun against five selected strains of diarrhoeagenic E. coli (DEC) were evaluated by the co-culture method. Results: Both drying methods significantly affected carotenoids and phenolic compounds. The Ogi slurry had viable LAB only for 10 days after which, there was a succession of fungi and yeast. Omidun showed 2 log10cfu/ml reduction of LAB count each week and the freeze-dried Ogi showed progressive reduction in viability. Refrigerated Omidun has little viable LAB, while higher viability was seen in modified Omidun (≥2 log cfu/ml) than normal Omidun. Modified Omidun intervention led to 2-4 log reduction in diarrhoeagenic E. coli strains and total inactivation of shigella-toxin producing E. coli H66D strain in co-culture. Conclusions: The consumption of Ogi should be within 10 days of milling using modified Omidun. There are practical potentials of consumption of Omidun in destroying E. coli strains implicated in diarrhea. Keywords: Ogi, Omidun, lactic acid bacteria, diarrhoeagenic Escherichia coli strains, Viability.

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