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Fulltext Engineering the production of major catechins by Escherichia coli carrying metabolite genes of Camellia sinensis

Umar KM, Abdulkarim SM, Radu S, Abdul Hamid A, Saari N

ScientificWorldJournal, 2012;2012:529031.
PMID: 22645428 DOI: 10.1100/2012/529031

A mimicked biosynthetic pathway of catechin metabolite genes from C. sinensis, consisting of flavanone 3 hydroxylase (F3H), dihydroflavonol reductase (DFR), and leucoanthocyanidin reductase (LCR), was designed and arranged in two sets of constructs: (a) single promoter in front of F3H and ribosome-binding sequences both in front of DFR and LCR; (b) three different promoters with each in the front of the three genes and ribosome-binding sequences at appropriate positions. Recombinant E. coli BL (DE3) harbouring the constructs were cultivated for 65 h at 26 °C in M9 medium consisting of 40 g/L glucose, 1 mM IPTG, and 3 mM eriodictyol. Compounds produced were extracted in ethyl acetate in alkaline conditions after 1 h at room temperature and identified by HPLC. Two of the four major catechins, namely, (-)-epicatechin (0.01) and (-)-epicatechin gallate (0.36 mg/L), and two other types ((+)-catechin hydrate (0.13 mg/L) and (-)-catechin gallate (0.04 mg/L)) were successfully produced.

Matched MeSH terms: Escherichia coli/genetics*; Escherichia coli/metabolism*
Phytochemical and biological evaluation of Defatted seeds of Jatropha Curcas

Ul Haq MN, Wazir SM, Ullah F, Khan RA, Shah MS, Khatak A

Sains Malaysiana, 2016;45:1435-1442.

In this study, the antimicrobial, antioxidant, phytotoxic and phytochemical properties of defatted seeds of Jatropha curcas were evaluated. A crude methanolic extract of defatted seeds was tested against three fungal strains - Aspergillus niger, Aspergillus flavus and Aspergillus fumigatus - and five bacteria: Escherichia coli and Klebsiella pneumoniae (Gram negative) and Micrococcus luteus, Bacillus subtilis and Staphylococcus aureus (Gram positive). The methanolic extract was diluted in dimethylsulfoxide to final concentrations of 1, 2, 3, 4 and 5 mg/10 mL. The largest zones of inhibition against K. pneumoniae, M. luteus and B. subtilis were achieved using the concentration of 5 mg/10 mL. The concentration of 1 mg/10 mL was most effective against S. aureus and E. coli. In a 1, 1-diphenyl-2-picrylahydrazyl (DPPH) radical scavenging assay, the 5 mg/10 mL concentration of the Jatropha seed extract showed the strongest activity. Higher concentrations of the Jatropha seed extract (10 mg/50 mL and 5 mg/50 mL) significantly inhibited the germination of radish seeds and had negative effects on radish seedling relative water content, shoot length, root length, seedling fresh weight and seedling dry weight (p<0.05). Phytochemical analyses of the defatted seeds detected alkaloids (7.3%), flavonoids (0.39%) and soluble phenolics (mg gallic acid equivalents/g extract). Based on these results, it was inferred that J. curcas seeds contain active ingredients that are effective against pathogenic microbes and therefore could be used to formulate drugs to treat various diseases.

Matched MeSH terms: Escherichia coli
Comparative study on microbial removal in immersed membrane filtration (IMF) with and without powdered activated carbon (PAC)

Ujang Z, Au YL, Nagaoka H

Water Sci Technol, 2002;46(9):109-15.
PMID: 12448459

This paper describes an investigation on the effect of microbial removal using IMF for high quality drinking water production. The comparison of IMF and IMF-PAC configuration was carried out in the study to highlight the importance of PAC in the system. The specific objective of this study was to study the effect of PAC adsorption in the IMF-PAC system particularly in removing microbial substances from contaminated raw water. A bench scale IMF-PAC configuration using a flat sheet microfiltration membrane was set up for experimental purposes. Experimentally, the result has shown high removal of microbial substances with the IMF-PAC system compared to IMF. The result of E. coli removal achieved was below the detectable level due to the microbial size, which is bigger than membrane pore size. The addition of PAC has shown a direct effect on total microbial removal. The adsorption of microbial onto PAC surfaces reduced the amount of smaller microbial present in permeate samples. As a conclusion, the configuration of IMF is a promising separation process in removing microbial substances, especially when the system is combined with PAC.

Matched MeSH terms: Escherichia coli/isolation & purification
In vitro assessment of pathogenicity and virulence encoding gene profiles of avian pathogenic Escherichia coli strains associated with colibacillosis in chickens

Ugwu IC, Lee-Ching L, Ugwu CC, Okoye JOA, Chah KF

Iran J Vet Res, 2020;21(3):180-187.
PMID: 33178295

Background: Avian pathogenic Escherichia coli (APEC) strains have been associated with various disease conditions in avian species due to virulence attributes associated with the organism.
Aims: This study was carried out to determine the in vitro pathogenic characteristics and virulence encoding genes found in E. coli strains associated with colibacillosis in chickens.
Methods: Fifty-two stock cultures of E. coli strains isolated from chickens diagnosed of colibacillosis were tested for their ability to produce haemolysis on blood agar and take up Congo red dye. Molecular characterization was carried out by polymerase chain reaction (PCR) amplification of virulence encoding genes associated with APEC.
Results: Eleven (22%) and 41 (71%) were positive for haemolysis on 5% sheep red blood agar and Congo red agar, respectively. Nine virulence-associated genes were detected as follows: FimH (96%), csgA (52%), iss (48%), iut (33%), tsh (21%), cva (15%), kpsII (10%), pap (2%), and felA (2%).
Conclusion: The APEC strains exhibited virulence properties and harbored virulence encoding genes which could be a threat to the poultry population and public health. The putative virulence genes were diverse and different in almost all isolate implying that pathogenesis was multi-factorial and the infection was multi-faceted which could be a source of concern in the detection and control of APEC infections.

Matched MeSH terms: Escherichia coli; Escherichia coli Infections; Adhesins, Escherichia coli; Escherichia coli Proteins
Fulltext Composition and Bioactivity of Essential Oil from Citrus grandis (L.) Osbeck 'Mato Peiyu' Leaf

Tsai ML, Lin CD, Khoo KA, Wang MY, Kuan TK, Lin WC, et al.

Molecules, 2017 Dec 05;22(12).
PMID: 29206180 DOI: 10.3390/molecules22122154

'Mato Peiyu' pomelo (Citrus grandis (L.) Osbeck 'Mato Peiyu') leaves from pruning are currently an agricultural waste. The aim of this study was to isolate essential oils from these leaves through steam distillation (SD) and solvent-free microwave extraction (SFME) and to evaluate their applicability to skin care by analyzing their antimicrobial, antioxidant (diphenyl-1-picrylhydrazyl scavenging assay, β-carotene/linoleic acid assay, and nitric oxide scavenging assay), anti-inflammatory (5-lipoxygenase inhibition assay), and antityrosinase activities. The gas chromatography-mass spectrometry results indicated that the main components of 'Mato Peiyu' leaf essential oils were citronellal and citronellol, with a total percentage of 50.71% and 59.82% for SD and SFME, respectively. The highest bioactivity among all assays was obtained for 5-lipoxygenase inhibition, with an IC50 value of 0.034% (v/v). The MIC90 of the antimicrobial activity of essential oils against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Candida albicans ranged from 0.086% to 0.121% (v/v). Citronellal and citronellol were the main contributors, accounting for at least 54.58% of the essential oil's bioactivity. This paper is the first to report the compositions and bioactivities of 'Mato Peiyu' leaf essential oil, and the results imply that the pomelo leaf essential oil may be applied in skin care.

Matched MeSH terms: Escherichia coli/drug effects; Escherichia coli/growth & development
Cloning of glyceraldehyde-3-phosphate dehydrogenase from an Antarctic psychrophilic bacterium by inverse and splinkerette PCR

Too WC, Liew YC, Few LL

J Basic Microbiol, 2008 Oct;48(5):430-5.
PMID: 18759222 DOI: 10.1002/jobm.200800008

Psychrophiles are organisms that thrive in cold environments. One of the strategies for their cold adaptation is the ability to synthesize cold-adapted enzymes. These enzymes usually display higher catalytic efficiency and thermolability at lower temperatures compared to their mesophilic and thermophilic counterparts. In this work, a psychrophilic bacterium codenamed pi9 was selected for the cloning of the gene encoding glyceraldehyde-3-phosphate dehydrogenase (GAPDH), an enzyme in the glycolytic pathway. Here, the cloning of an 1,113 bp fragment of GAPDH gene which covers the 1,002 bp open reading frame by using multiple PCR steps is described. The partial sequence of this gene was PCR amplified by using degenerate primers followed by the cloning of the flanking sequences by inverse and splinkerette PCR techniques. The success in cloning the GAPDH gene by PCR has bypassed the more time consuming genomic library construction and screening method. The full length GAPDH protein was subsequently expressed in E. coli, purified as His-tag protein and confirmed to be catalytically active. This work demonstrated the use of multiple PCR techniques to clone a gene based solely on sequence comparison. It also laid the foundation for further biochemical and structural characterizations of GAPDH from a psychrophilic bacterium by providing a highly purified recombinant protein sample.

Matched MeSH terms: Escherichia coli/enzymology; Escherichia coli/genetics
Enhancing organic matter productivity in microalgal-bacterial biofilm using novel bio-coating

Tong CY, Honda K, Derek CJC

Sci Total Environ, 2024 Jan 01;906:167576.
PMID: 37804964 DOI: 10.1016/j.scitotenv.2023.167576

Research on renewable energy from microalgae has led to a growing interest in porous substrate photobioreactors, but their widespread adoption is currently limited to pure microalgal biofilm cultures. The behavior of microalgal-bacterial biofilms immobilized on microporous substrates remains as a research challenge, particularly in uncovering their mutualistic interactions in environment enriched with dissolved organic matter. Therefore, this study established a novel culture platform by introducing microalgal-derived bio-coating that preconditioned hydrophilic polyvinylidene fluoride membranes for the microalgal-bacterial biofilm growth of freshwater microalgae, Chlorella vulgaris ESP 31 and marine microalgae, Cylindrotheca fusiformis with bacteria, Escherichia coli. In the attached co-culture mode, the bio-coating we proposed demonstrated the ability to enhance microalgal growth for both studied species by a range of 2.5 % to 19 % starting from day 10 onwards. Additionally, when compared to co-culture on uncoated membranes, the bio-coating exhibited a significant bacterial growth promotion effect, increasing bacterial growth by at least 2.35 times for the C. vulgaris-E. coli co-culture after an initial adaptation phase. A significant increase of at least 72 % in intracellular biochemical compounds (including chlorophyll, polysaccharides, proteins, and lipids) was observed within just five days, primarily due to the high concentration of pre-coated organic matter, mainly sourced from the internal organic matter (IOM) of C. fusiformis. Higher accumulation of organic compounds in the bio-coating indirectly triggers a competition between microalgae and bacteria which potentially stimulate the production of additional intra-/extra-organic substances as a defensive response. In short, insight gained from this study may represent a paradigm shift in the ways that symbiotic interactions are promoted to increase the yield of specific bio-compounds with the presence of bio-coating.

Matched MeSH terms: Escherichia coli
Role of coaggregation in the pathogenicity and prolonged colonisation of Vibrio cholerae

Toh YS, Yeoh SL, Yap IKS, Teh CSJ, Win TT, Thong KL, et al.

Med Microbiol Immunol, 2019 Dec;208(6):793-809.
PMID: 31263955 DOI: 10.1007/s00430-019-00628-3

Cholera is an acute diarrheal illness caused by the Gram-negative bacterium Vibrio cholerae. The pathogen is known for its ability to form biofilm that confers protection against harsh environmental condition and as part of the colonisation process during infection. Coaggregation is a process that facilitates the formation of biofilm. In a preliminary in vitro study, high coaggregation index and biofilm production were found between V. cholerae with human commensals namely Escherichia coli and Enterobacter cloacae. Building upon these results, the effects of coaggregation were further evaluated using adult BALB/c mouse model. The animal study showed no significant differences in mortality and fluid accumulation ratio between treatment groups infected with V. cholerae alone and those infected with coaggregation partnership (V. cholerae with E. coli or V. cholerae with E. cloacae). However, mild inflammation was detected in both partnering pairs. Higher density of V. cholerae was recovered from faecal samples of mice co-infected with E. coli and V. cholerae in comparison with other groups at 24 h post-infection. This partnership also elicited slightly higher levels of interleukin-5 (IL-5) and interleukin-10 (IL-10). Nonetheless, the involvement of autoinducer-2 (AI-2) as the signalling molecules in quorum sensing system is not evident in this study. Since E. coli is one of the common commensals, our result may suggest the involvement of commensals in cholera development.

Matched MeSH terms: Escherichia coli/growth & development
Phenotypic evaluation of ESBL- and carbapenemaseproducing Escherichia coli and Klebsiella pneumoniae from a teaching hospital in the Philippines

Tiongco RE, Arceo E, Dizon D, Navarro A, Rivera N, Salita C, et al.

Trop Biomed, 2018 Dec 01;35(4):1064-1074.
PMID: 33601853

Antimicrobial resistance is a worldwide public health concern. Rise in the number of antimicrobial resistant organisms, such as extended spectrum β-lactamase- (ESBL) and carbapenemase-producing Escherichia coli and Klebsiella pneumoniae, continue to burden millions of people worldwide. E. coli and K. pneumoniae were isolated and collected for four months from a teaching hospital in the Philippines. All isolates were subjected to ESBL and carbapenemase testing using the double disk synergy test and modified Hodge test, respectively. Their pattern of resistance among different classes of antimicrobial agents was also investigated using the Kirby-Bauer disk diffusion test. Among the 32 clinical isolates tested, 28.1% were positive for ESBL production and 6.3% were positive for carbapenemase production. Species-specific classification showed that E. coli (44.4%) has the highest rate of ESBL production whereas both E. coli (5.6%) and K. pneumoniae (7.1%) showed almost similar rates of carbapenemase production. Antimicrobial resistance pattern of drug resistant isolates showed that all organisms were resistant to ampicillin, and majority showed resistance towards ciprofloxacin, cefotaxime, ceftriaxone, and sulfamethoxazole/trimethoprim. ESBL production is seen highest among E. coli isolates while similar rates of carbapenemase production was observed to both E. coli and K. pneumoniae isolates. Overall, antimicrobial resistance continues to rise and poses a huge threat in public health worldwide. Efforts should be made in developing rapid tests for antimicrobial resistance and to search for effective treatment from infections caused by multidrug resistant organisms.

Matched MeSH terms: Escherichia coli
Fulltext Serum from Nipah Virus Patients Recognises Recombinant Viral Proteins Produced in Escherichia coli

Tiong V, Lam CW, Phoon WH, AbuBakar S, Chang LY

Jpn J Infect Dis, 2017 Jan 24;70(1):26-31.
PMID: 27169942 DOI: 10.7883/yoken.JJID.2015.501

The genes for Nipah virus (NiV) proteins were amplified from viral RNA, cloned into the plasmid pTriEx-3 Hygro, expressed, and purified using immobilized metal affinity chromatography. The recombinant N, F, and G NiV proteins (rNiV-N, rNiV-F, and rNiV-G), were successfully expressed in Escherichia coli and purified with a yield of 4, 16, and 4 mg/L, respectively. All 3 recombinant viral proteins reacted with all 19 samples of NiV-positive human sera. The rNiV-N and rNiV-G proteins were the most immunogenic. The recombinant viral proteins did not react with any of the 12 NiV-negative sera. However, serum from a patient with a late-onset relapsing NiV infection complication was found to be primarily reactive to rNiV-G only. Additionally, there is a distinctive variation in the profile of antigen-reactive bands between the sample from a case of relapsing NiV encephalitis and that of acute NiV infection. The overall findings of this study suggest that the recombinant viral proteins have the potential to be developed further for use in the detection of NiV infection, and continuous biosurveillance of NiV infection in resource-limited settings.

Matched MeSH terms: Escherichia coli/genetics; Escherichia coli/metabolism
Fulltext Inhibitory potency of 8-methoxypsoralen on cytochrome P450 2A6 (CYP2A6) allelic variants CYP2A6 15, CYP2A6 16, CYP2A6 21 and CYP2A6 22: differential susceptibility due to different sequence locations of the mutations

Tiong KH, Mohammed Yunus NA, Yiap BC, Tan EL, Ismail R, Ong CE

PLoS One, 2014;9(1):e86230.
PMID: 24475091 DOI: 10.1371/journal.pone.0086230

Human cytochrome P450 2A6 (CYP2A6) is a highly polymorphic isoform of CYP2A subfamily. Our previous kinetic study on four CYP2A6 allelic variants (CYP2A6 15, CYP2A6 16, CYP2A6 21 and CYP2A6 22) have unveiled the functional significance of sequence mutations in these variants on coumarin 7-hydroxylation activity. In the present study, we further explored the ability of a typical CYP2A6 inhibitor, 8-methoxypsoralen (8-MOP), in inhibition of these alleles and we hypothesized that translational mutations in these variants are likely to give impact on 8-MOP inhibitory potency. The CYP2A6 variant and the wild type proteins were subjected to 8-MOP inhibition to yield IC50 values. In general, a similar trend of change in the IC50 and Km values was noted among the four mutants towards coumarin oxidation. With the exception of CYP2A6 16, differences in IC50 values were highly significant which implied compromised interaction of the mutants with 8-MOP. Molecular models of CYP2A6 were subsequently constructed and ligand-docking experiments were performed to rationalize experimental data. Our docking study has shown that mutations have induced enlargement of the active site volume in all mutants with the exception of CYP2A6 16. Furthermore, loss of hydrogen bond between 8-MOP and active site residue Asn297 was evidenced in all mutants. Our data indicate that the structural changes elicited by the sequence mutations could affect 8-MOP binding to yield differential enzymatic activities in the mutant CYP2A6 proteins.

Matched MeSH terms: Escherichia coli
Fulltext Functional characterization of cytochrome P450 2A6 allelic variants CYP2A6*15, CYP2A6*16, CYP2A6*21, and CYP2A6*22

Tiong KH, Yiap BC, Tan EL, Ismail R, Ong CE

Drug Metab. Dispos., 2010 May;38(5):745-51.
PMID: 20139165 DOI: 10.1124/dmd.109.031054

Variation in CYP2A6 levels and activity can be attributed to genetic polymorphism and, thus, functional characterization of allelic variants is necessary to define the importance of CYP2A6 polymorphism in humans. The aim of the present study was to investigate the reported alleles CYP2A6*15, CYP2A6*16, CYP2A6*21, and CYP2A6*22, in terms of the functional consequences of their mutations on the enzyme catalytic activity. With use of the wild-type CYP2A6 cDNA as template, site-directed mutagenesis was performed to introduce nucleotide changes encoding K194E substitution in CYP2A6*15, R203S substitution in CYP2A6*16, K476R substitution in CYP2A6*21, and concurrent D158E and L160I substitutions in CYP2A6*22. Upon sequence verification, the CYP2A6 wild-type and mutant constructs were individually coexpressed with NADPH-cytochrome P450 reductase in Escherichia coli. A kinetic study using a coumarin 7-hydroxylase assay indicated that CYP2A6*15 exhibited higher V(max) than the wild type, whereas all mutant constructs, except for variant CYP2A6*16, exhibited higher K(m) values. Analysis of the V(max)/K(m) ratio revealed that all mutants demonstrated 0.85- to 1.05-fold differences from the wild type, with the exception of variant CYP2A6*22, which only portrayed 39% of the wild-type intrinsic clearance. These data suggested that individuals carrying the CYP2A6*22 allele are likely to have lower metabolism of CYP2A6 substrate than individuals expressing CYP2A6*15, CYP2A6*16, CYP2A6*21, and the wild type.

Matched MeSH terms: Escherichia coli/genetics; Escherichia coli/metabolism
Prevalence of AmpC beta-lactamase and extended spectrum beta-lactamase co-producer in Escherichia coli and Klebsiella species in a teaching hospital

Ting SMV, Ismail Z, Hanafiah A

Malays J Pathol, 2024 Apr;46(1):79-89.
PMID: 38682847

INTRODUCTION: Beta-lactamase producing bacterial infection has been on surge due to selection pressure and injudicious antibiotics usage. Organisms that co-produced more than one beta lactamase enzyme posed diagnostic challenges which may result in inadequate treatment. To date, there is no standardised guideline offering phenotypic detection of AmpC β-lactamase. The purpose of this study was to determine the prevalence of ESBLs, AmpC β-lactamase and co-producer organisms in a teaching hospital.
MATERIALS AND METHODS: Three hundred and four isolates of E. coli and Klebsiella sp. had been selected via convenient sampling. These isolates were identified using conventional laboratory methods and their antimicrobial susceptibilities were determined using disc diffusion method. Those isolates were then proceeded with ESBL confirmatory test, cloxacillin-containing Muller Hinton confirmatory test, modified double disk synergy test and AmpC disk test.
RESULTS: Out of 304 isolates, 159 isolates were E. coli and 145 were Klebsiella sp. The prevalence of organisms which co-produced AmpC β-lactamase and ESBL enzymes were 3.0%. Besides that, 39 cefoxitin resistant and three cefoxitin susceptible isolates (13.8%) were proven to produce AmpC β-lactamase through AmpC disk test. Through the CLSI confirmatory test, 252 (82.9%) isolates were identified as ESBLs producers and the prevalence increased slightly when cloxacillin-containing Muller Hinton were used. Only three ESBLs positive organisms were positive for modified double disk synergy test.
CONCLUSION: Distinguishing between AmpC β-lactamase and ESBL-producing organisms has epidemiological significance as well as therapeutic importance. Moreover, AmpC β-lactamase and ESBLs co-producing organisms can lead to false negative ESBL confirmatory test. Therefore, knowing the local prevalence can guide the clinician in navigating the treatment.

Matched MeSH terms: Escherichia coli Infections/microbiology; Escherichia coli Infections/epidemiology
Fulltext Simultaneous detection of methicillin-resistant Staphylococcus aureus, Acinetobacter baumannii, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa by multiplex PCR

Thong KL, Lai MY, Teh C SJ, Chua KH

Trop Biomed, 2011 Apr;28(1):21-31.
PMID: 21602765 MyJurnal

A PCR-based assay that can simultaneously detect and differentiate five different types of nosocomial bacterial pathogens was developed. Six pairs of selected primers targeting femA (132 bp) and mecA (310 bp) of methicillin-resistant Staphylococcus aureus, gltA (722 bp) of Acinetobacter baumannii, phoA (903 bp) of Escherichia coli, mdh (364 bp) of Klebsiella pneumoniae and oprL (504 bp) of Pseudomonas aeruginosa were used in this study. The conditions were optimized for the multiplex PCR to ensure specific amplification of the selected targets. Sensitivity and specificity tests were also carried out using a blind test approach on 50 bacterial cultures and resulted in 100% for both positive and negative predictive values.

Matched MeSH terms: Escherichia coli/isolation & purification
Fulltext Whole-Genome Sequence of Endophytic Plant Growth-Promoting Escherichia coli USML2

Tharek M, Sim KS, Khairuddin D, Ghazali AH, Najimudin N

Genome Announc, 2017 May 11;5(19).
PMID: 28495774 DOI: 10.1128/genomeA.00305-17

Escherichia coli strain USML2 was originally isolated from the inner leaf tissues of surface-sterilized phytopathogenic-free oil palm (Elaeis guineensis Jacq.). We present here the whole-genome sequence of this plant-endophytic strain. The genome consists of a single circular chromosome of 4,502,758 bp, 4,315 predicted coding sequences, and a G+C content of 50.8%.

Matched MeSH terms: Escherichia coli
Fulltext Effect of different cloning strategies in pET-28a on solubility and functionality of a staphylococcal phage endolysin

Tham HY, Song AA, Yusoff K, Tan GH

Biotechniques, 2020 09;69(3):161-170.
PMID: 32787565 DOI: 10.2144/btn-2020-0034

Endolysins have been studied intensively as an alternative to antibiotics. In this study, endolysin derived from a phage which infects methicillin-resistant Staphylococcus aureus (MRSA) was cloned and expressed in Escherichia coli pET28a. Initially, the endolysin was cloned using BamHI/XhoI, resulting in expression of a recombinant endolysin which was expressed in inclusion bodies. While solubilization was successful, the protein remained nonfunctional. Recloning the endolysin using NcoI/XhoI resulted in expression of soluble and functional proteins at 18°C. The endolysin was able to form halo zones on MRSA plates and showed a reduction in turbidity of MRSA growth. Therefore, cloning strategies should be chosen carefully even in an established expression system as they could greatly affect the functionality of the expressed protein.

Matched MeSH terms: Escherichia coli/genetics
Fulltext Production and purification of polymerization-competent HIV-1 capsid protein p24 (CA) in NiCo21(DE3) Escherichia coli

Teow SY, Mualif SA, Omar TC, Wei CY, Yusoff NM, Ali SA

BMC Biotechnol, 2013;13:107.
PMID: 24304876 DOI: 10.1186/1472-6750-13-107

HIV genome is packaged and organized in a conical capsid, which is made up of ~1,500 copies of the viral capsid protein p24 (CA). Being a primary structural component and due to its critical roles in both late and early stages of the HIV replication cycle, CA has attracted increased interest as a drug discovery target in recent years. Drug discovery studies require large amounts of highly pure and biologically active protein. It is therefore desirable to establish a simple and reproducible process for efficient production of HIV-1 CA.

Matched MeSH terms: Escherichia coli/genetics*; Escherichia coli/metabolism
Characterization of Thiomonas delicata arsenite oxidase expressed in Escherichia coli

Teoh WK, Salleh FM, Shahir S

3 Biotech, 2017 Jun;7(2):97.
PMID: 28560637 DOI: 10.1007/s13205-017-0740-7

Microbial arsenite oxidation is an essential biogeochemical process whereby more toxic arsenite is oxidized to the less toxic arsenate. Thiomonas strains represent an important arsenite oxidizer found ubiquitous in acid mine drainage. In the present study, the arsenite oxidase gene (aioBA) was cloned from Thiomonas delicata DSM 16361, expressed heterologously in E. coli and purified to homogeneity. The purified recombinant Aio consisted of two subunits with the respective molecular weights of 91 and 21 kDa according to SDS-PAGE. Aio catalysis was optimum at pH 5.5 and 50-55 °C. Aio exhibited stability under acidic conditions (pH 2.5-6). The V max and K m values of the enzyme were found to be 4 µmol min(-1) mg(-1) and 14.2 µM, respectively. SDS and Triton X-100 were found to inhibit the enzyme activity. The homology model of Aio showed correlation with the acidophilic adaptation of the enzyme. This is the first characterization studies of Aio from a species belonging to the Thiomonas genus. The arsenite oxidase was found to be among the acid-tolerant Aio reported to date and has the potential to be used for biosensor and bioremediation applications in acidic environments.

Matched MeSH terms: Escherichia coli
Fulltext Immunogenicity of recombinant BCG-based vaccine expressing the 22 kDa of serine repeat antigen (SE22) of Plasmodium falciparum

Teo WH, Nurul AA, Norazmi MN

Trop Biomed, 2012 Jun;29(2):239-53.
PMID: 22735846 MyJurnal

The Plasmodium falciparum serine repeat antigen (SERA) is one of the promising blood-stage malarial vaccine candidates. In this study, recombinant Mycobacterium bovis bacille Calmette-Guerin (rBCG) expressing the 22 kDa protein (SE22) from the 47 kDa Nterminal domain of serine repeat antigen (SERA), generated in favour of mycobacterium codon usage, elicited specific immune response in BALB/c mice with a mixed Th1/Th2 profile. Immunized sera containing high levels of specific IgG1 and IgG2a against the epitope (as determined by ELISA) were reactive with fixed P. falciparum merozoites as demonstrated by indirect immunofluorescence assay (IFA). Furthermore, the lymphocyte proliferative response to SE22 antigen from rBCG-immunized mice was higher than that of controls. The expression of intracellular cytokines (IL-2, IL-4 and IFNγ) in CD4+- and CD8+-cells was also enhanced following in-vitro stimulation with SE22. These findings indicate that a rBCG-based vaccine candidate expressing a blood-stage antigen of P. falciparum could enhance both humoral and cellular immune responses, thus paving the way for the rational use of rBCG as a vaccine candidate against malaria.

Matched MeSH terms: Escherichia coli/genetics; Escherichia coli/metabolism
Fulltext Bioactive Compounds from the Bornean Endemic Plant Goniothalamus longistipetes

Teo SP, Bhakta S, Stapleton P, Gibbons S

Antibiotics (Basel), 2020 Dec 16;9(12).
PMID: 33339285 DOI: 10.3390/antibiotics9120913

The present study aimed to screen plants for bioactive compounds with potential antibacterial activities. In our efforts to evaluate plants from Borneo, we isolated and elucidated the structures of four natural products from the bioactive fraction of a chloroform extract of Goniothalamus longistipetes using various chromatographic and spectroscopic techniques. The bioactive compounds were identified as a known styryllactone, (+)-altholactone ((2S,3R,3aS,7aS)-3-hydroxy-2-phenyl-2,3,3a,7a-tetrahydrobenzo-5(4H)-5-one) (1), a new styryllactone, (2S,3R,3aS,7aS)-3-hydroxy-2-phenyl-2,3,3a,7a-tetrahydrobenzo-5(4H)-5-one) (2) as well as a new alkaloid, 2,6-dimethoxyisonicotinaldehyde (3) and a new alkenyl-5-hydroxyl-phenyl benzoic acid (4). 1 and 4 showed broad-spectrum anti-bacterial activities against Gram-positive and Gram-negative bacteria as well as acid-fast model selected for this study. Compound 2 only demonstrated activities against Gram-positive bacteria whilst 3 displayed selective inhibitory activities against Gram-positive bacterial strains. Additionally, their mechanisms of anti-bacterial action were also investigated. Using Mycobacterium smegmatis as a fast-growing model of tubercle bacilli, compounds 1, 2 and 4 demonstrated inhibitory activities against whole-cell drug efflux and biofilm formation; two key intrinsic mechanisms of antibiotic resistance. Interestingly, the amphiphilic compound 4 exhibited inhibitory activity against the conjugation of plasmid pKM101 in Escherichia coli using a plate conjugation assay. Plasmid conjugation is a mechanism by which Gram-positive and Gram-negative-bacteria acquire drug resistance and virulence. These results indicated that bioactive compounds isolated from Goniothalamus longistipetes can be potential candidates as 'hits' for further optimisation.

Matched MeSH terms: Escherichia coli

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