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Fulltext Antigenic membrane proteins of virulent variant of Entamoeba histolytica HM-1:IMSS

Kumarasamy G, Abdus Sani AA, Olivos-García A, Noordin R, Othman N

Pathog Glob Health, 2020 09;114(6):333-342.
PMID: 32536281 DOI: 10.1080/20477724.2020.1780402

Amoebiasis, caused by Entamoeba histolytica, is one of the leading parasitic infections in the world. This study was aimed at profiling antigenic membrane proteins of a virulent variant of E. histolytica HM-1:IMSS. The membrane proteins were extracted using ProteoExtract® kit (Merck, Darmstadt, Germany) or conventional method, separated using OFFGEL 3100 fractionator (Agilent Technologies, Santa Clara, California), followed by SDS-PAGE and Western blot analysis. Selected antigenic membrane proteins were identified using LC-ESI-MS/MS. Subsequently, the proteins were classified according to their biological processes and predictions were made on membrane and membrane-associated proteins. When the proteins were probed with pooled sera from amoebic liver abscess (ALA) patients, 10 and 15 antigenic proteins with molecular weights 25 to 200 kDa were identified using the ProteoExtract® kit and conventional method, respectively. LC-ESI-MS/MS identified 13 antigenic proteins, and both extraction methods predicted six of them as membrane and membrane-associated proteins. The topmost biological processes which comprised of six proteins were involved in cellular processes.. These antigenic membrane proteins merit further investigations as potential candidates for vaccine studies.

Matched MeSH terms: Membrane Proteins/immunology*; Protozoan Proteins/immunology*
Fulltext The Role of the Z-DNA Binding Domain in Innate Immunity and Stress Granules

Chiang C, Li Y, Ng SK

Front Immunol, 2020;11:625504.
PMID: 33613567 DOI: 10.3389/fimmu.2020.625504

Both DNA and RNA can maintain left-handed double helical Z-conformation under physiological condition, but only when stabilized by Z-DNA binding domain (ZDBD). After initial discovery in RNA editing enzyme ADAR1, ZDBD has also been described in pathogen-sensing proteins ZBP1 and PKZ in host, as well as virulence proteins E3L and ORF112 in viruses. The host-virus antagonism immediately highlights the importance of ZDBD in antiviral innate immunity. Furthermore, Z-RNA binding has been shown to be responsible for the localization of these ZDBD-containing proteins to cytoplasmic stress granules that play central role in coordinating cellular response to stresses. This review sought to consolidate current understanding of Z-RNA sensing in innate immunity and implore possible roles of Z-RNA binding within cytoplasmic stress granules.

Matched MeSH terms: DNA-Binding Proteins/immunology*; Viral Proteins/immunology*; RNA-Binding Proteins/immunology*
Immunoproteomic analysis of antibody response to cell wall-associated proteins of Candida tropicalis

Lee PY, Gam LH, Yong VC, Rosli R, Ng KP, Chong PP

J Appl Microbiol, 2014 Sep;117(3):854-65.
PMID: 24909754 DOI: 10.1111/jam.12562

This study was conducted to identify antigenic proteins of Candida tropicalis that are targeted by the host immune system.

Matched MeSH terms: Fungal Proteins/immunology*; Membrane Proteins/immunology*; Recombinant Proteins/immunology
Recombinant proteins in the diagnosis of toxoplasmosis

Kotresha D, Noordin R

APMIS, 2010 Aug;118(8):529-42.
PMID: 20666734 DOI: 10.1111/j.1600-0463.2010.02629.x

Toxoplasma gondii is an important human pathogen with a worldwide distribution. It is primarily of medical importance for pregnant women and immunocompromised patients. Primary infection of the former is often associated with fetal infection, which can lead to abortion or severe neonatal malformation. Immunocompromised patients are at risk of contracting the severe form of the disease that may be fatal. Thus, detection of T. gondii infection with high sensitivity and specificity is crucial in the management of the disease. Toxoplasmosis is generally diagnosed by demonstrating specific immunoglobulin M (IgM) and IgG antibodies to toxoplasma antigens in the patient's serum sample. Most of the commercially available tests use T. gondii native antigens and display wide variations in test accuracy. Recombinant antigens have great potential as diagnostic reagents for use in assays to detect toxoplasmosis. Thus in this review, we address recent advances in the use of Toxoplasma recombinant proteins for serodiagnosis of toxoplasmosis.

Matched MeSH terms: Membrane Proteins/immunology; Recombinant Proteins/immunology; Protozoan Proteins/immunology
Induction and characterization of heat shock proteins of Salmonella typhi and their reactivity with sera from patients with typhoid fever

Tang SW, Abubakar S, Devi S, Puthucheary S, Pang T

Infect Immun, 1997 Jul;65(7):2983-6.
PMID: 9199477

The heat shock protein (HSP) response of Salmonella typhi following exposure to elevated growth temperatures was studied. Three major proteins with molecular sizes of 58, 68, and 88 kDa were abundantly expressed when S. typhi cells were shifted from 37 to 45 degrees C and to 55 degrees C. These proteins were also constitutively expressed at 37 degrees C. Western blotting and immunoprecipitation studies with anti-HSP monoclonal antibodies revealed that the 58- and 68-kDa proteins were analogous to the GroEL and DnaK proteins, respectively, of Escherichia coli. These HSPs are also abundantly present in the outer membrane fraction of disrupted cells and, to a lesser extent, in the cytosol. Immunoblotting experiments with sera from patients with a culture-positive diagnosis of typhoid fever showed the presence of antibodies to these HSPs. Nine of twelve sera reacted with the 58-, 68-, and 88-kDa proteins, while three sera reacted only with the 68- and 88-kDa proteins. All 10 sera from healthy individuals showed no binding to these HSPs. In light of the well-documented roles of HSPs in the pathogenesis of microbial infections and as immunodominant antigens, these findings may be relevant for a better understanding of disease processes and for the future development of diagnostic and preventive strategies.

Matched MeSH terms: Bacterial Proteins/immunology; Heat-Shock Proteins/immunology*; HSP70 Heat-Shock Proteins/immunology
Inclusion bodies of recombinant Epstein-Barr virus capsid antigen p18 as potential immobilized antigens in enzyme immunoassays for detection of nasopharyngeal carcinoma

Lim CS, Goh SL, Kariapper L, Krishnan G, Lim YY, Ng CC

Clin Chim Acta, 2015 Aug 25;448:206-10.
PMID: 26164385 DOI: 10.1016/j.cca.2015.07.008

Development of indirect enzyme-linked immunosorbent assays (ELISAs) often utilizes synthetic peptides or recombinant proteins from Escherichia coli as immobilized antigens. Because inclusion bodies (IBs) formed during recombinant protein expression in E. coli are commonly thought as misfolded aggregates, only refolded proteins from IBs are used to develop new or in-house diagnostic assays. However, the promising utilities of IBs as nanomaterials and immobilized enzymes as shown in recent studies have led us to explore the potential use of IBs of recombinant Epstein-Barr virus viral capsid antigen p18 (VCA p18) as immobilized antigens in ELISAs for serologic detection of nasopharyngeal carcinoma (NPC).

Matched MeSH terms: Recombinant Proteins/immunology; Capsid Proteins/immunology*; Immobilized Proteins/immunology*
Natural cryptic autoantibodies

Cheng HM

Autoimmunity, 1998;27(2):99-108.
PMID: 9583741
Matched MeSH terms: Blood Proteins/immunology
Fulltext Autoantibodies to survivin in the sera of patients with infiltrating ductal carcinoma of the breast

Al-Joudi FS, Iskandar ZA

Med J Malaysia, 2006 Aug;61(3):302-6.
PMID: 17240580 MyJurnal

Autoantibodies to survivin have been reported in lung cancers and in gastrointestinal cancers. A few reports have also described a low prevalence of autoantibodies to survivin and at low titres in the sera of breast cancer patients with no implications for their clinical usefulness. This study was designed to re-examine the prevalence and the clinical correlations of autoantibodies to the tumour protein survivin, in the sera of patients with infiltrating ductal carcinoma of the breast using an ELISA assay. In spite of the low prevalence of autoantibodies to survivin (7%, n = 57), their presence was associated with grade III tumours, with tumour sizes exceeding 10cm, with axillary lymph nodal involvement and with metastases. Moreover, all the autoantibody-positive cases were estrogen and progesterone receptors negative. Furthermore, all the autoantibody-positive cases expressed survivin with high scores.

Matched MeSH terms: Microtubule-Associated Proteins/immunology*; Neoplasm Proteins/immunology*
Expression and Purification of E2 Glycoprotein from Insect Cells (Sf9) for Use in Serology

Chua CL, Sam IC, Chan YF

Methods Mol Biol, 2016;1426:51-61.
PMID: 27233260 DOI: 10.1007/978-1-4939-3618-2_5

Chikungunya virus (CHIKV) is a mosquito-borne arbovirus which poses a major threat to global public health. Definitive CHIKV diagnosis is crucial, especially in distinguishing the disease from dengue virus, which co-circulates in endemic areas and shares the same mosquito vectors. Laboratory diagnosis is mainly based on serological or molecular approaches. The E2 glycoprotein is a good candidate for serological diagnosis since it is the immunodominant antigen during the course of infection, and reacts with seropositive CHIKV sera. In this chapter, we describe the generation of stable clone Sf9 (Spodoptera frugiperda) cells expressing secreted, soluble, and native recombinant CHIKV E2 glycoprotein. We use direct plasmid expression in insect cells, rather than the traditional technique of generating recombinant baculovirus. This recombinant protein is useful for serological diagnosis of CHIKV infection.

Matched MeSH terms: Recombinant Proteins/immunology; Viral Envelope Proteins/immunology*
Fulltext Antibody responses of dengue fever patients to dengue 2 (New Guinea C strain) viral proteins

AbuBakar S, Azmi A, Mohamed-Saad N, Shafee N, Chee HY

Malays J Pathol, 1997 Jun;19(1):41-51.
PMID: 10879241

The present study was undertaken to investigate the antibody responses of dengue fever (DF) patients to specific dengue virus proteins. Partially purified dengue 2 New Guinea C (NGC) strain virus was used as antigen. Under the present experimental protocols, it was observed that almost all DF patients' sera had detectable presence of antibodies which recognize the dengue 2 envelope (E) protein. The convalescent-phase sera especially had significant detectable IgG, IgM and IgE against the protein. In addition, IgGs specific against the NS1 dimer and PrM were also detected. Antibody against the core (C) protein, however, was not detectable in any of the DF patients' sera. The substantial presence of IgG against the PrM in the convalescent-phase sera, and the presence of IgE specific for the E, reflect the potential importance of these antibody responses in the pathogenesis of dengue.

Matched MeSH terms: Viral Envelope Proteins/immunology*; Viral Nonstructural Proteins/immunology*
Generation of a naïve human single chain variable fragment (scFv) library for the identification of monoclonal scFv against Salmonella Typhi Hemolysin E antigen

Lim BN, Chin CF, Choong YS, Ismail A, Lim TS

Toxicon, 2016 Jul;117:94-101.
PMID: 27090555 DOI: 10.1016/j.toxicon.2016.04.032

Antibody phage display is a useful tool for the isolation and identification of monoclonal antibodies. Naive antibody libraries are able to overcome the limitations associated with the traditional hybridoma method for monoclonal antibody generation. Antibody phage display is also a preferred method for antibody generation against toxins as it does not suffer from toxicity mediated complications. Here, we describe a naïve multi ethnic scFv antibody library generated via two-step cloning with an estimated diversity of 2 × 10(9). The antibody library was used to screen for monoclonal antibodies against Hemolysin E antigen, a pore forming toxin produced by Salmonella enterica serovar Typhi. A soluble monoclonal scFv antibody against the HlyE toxin (IgM scFv D7 anti-hlyE) was isolated from the library. This shows the value of the naïve library to generate antibodies against toxin targets in addition to the potential use of the library to isolate antibodies against other immunogenic targets.

Matched MeSH terms: Hemolysin Proteins/immunology*; Escherichia coli Proteins/immunology*
Fulltext In vitro invasion inhibition assay using antibodies against Plasmodium knowlesi Duffy binding protein alpha and apical membrane antigen protein 1 in human erythrocyte-adapted P. knowlesi A1-H.1 strain

Muh F, Lee SK, Hoque MR, Han JH, Park JH, Firdaus ER, et al.

Malar J, 2018 Jul 27;17(1):272.
PMID: 30049277 DOI: 10.1186/s12936-018-2420-4

BACKGROUND: The rapid process of malaria erythrocyte invasion involves ligand-receptor interactions. Inducing antibodies against specific ligands or receptors that abrogate the invasion process is a key challenge for blood stage vaccine development. However, few candidates were reported and remain to be validated for the discovery of new vaccine candidates in Plasmodium knowlesi.
METHODS: In order to investigate the efficacy of pre-clinical vaccine candidates in P. knowlesi-infected human cases, this study describes an in vitro invasion inhibition assay, using a P. knowlesi strain adapted to in vitro growth in human erythrocytes, PkA1-H.1. Recombinant proteins of P. knowlesi Duffy binding protein alpha (PkDBPα) and apical membrane antigen 1 (PkAMA1) were produced in Escherichia coli system and rabbit antibodies were generated from immune animals.
RESULTS: PkDBPα and PkAMA1 recombinant proteins were expressed as insoluble and produced as a functional refolded form for this study. Antibodies against PkDBPα and PkAMA1 specifically recognized recombinant proteins and native parasite proteins in schizont-stage parasites on the merozoite organelles. Single and combination of anti-PkDBPα and anti-PkAMA1 antibodies elicited strong growth inhibitory effects on the parasite in concentration-dependent manner. Meanwhile, IgG prevalence of PkDBPα and PkAMA1 were observed in 13.0 and 46.7% in human clinical patients, respectively.
CONCLUSION: These data provide support for the validation of in vitro growth inhibition assay using antibodies of DBPα and AMA1 in human-adapted P. knowlesi parasite PkA1-H.1 strain.

Matched MeSH terms: Membrane Proteins/immunology*; Protozoan Proteins/immunology*
DNA vaccination with a plasmid encoding LACK-TSA fusion against Leishmania major infection in BALB/c mice

Maspi N, Ghaffarifar F, Sharifi Z, Dalimi A, Khademi SZ

Malays J Pathol, 2017 Dec;39(3):267-275.
PMID: 29279589

Vaccination would be the most important strategy for the prevention and elimination of leishmaniasis. The aim of the present study was to compare the immune responses induced following DNA vaccination with LACK (Leishmania analogue of the receptor kinase C), TSA (Thiol-specific-antioxidant) genes alone or LACK-TSA fusion against cutaneous leishmaniasis (CL). Cellular and humoral immune responses were evaluated before and after challenge with Leishmania major (L. major). In addition, the mean lesion size was also measured from 3th week post-infection. All immunized mice showed a partial immunity characterized by higher interferon (IFN)-γ and Immunoglobulin G (IgG2a) levels compared to control groups (p<0.05). IFN-γ/ Interleukin (IL)-4 and IgG2a/IgG1 ratios demonstrated the highest IFN-γ and IgG2a levels in the group receiving LACK-TSA fusion. Mean lesion sizes reduced significantly in all immunized mice compared with control groups at 7th week post-infection (p<0.05). In addition, there was a significant reduction in mean lesion size of LACK-TSA and TSA groups than LACK group after challenge (p<0.05). In the present study, DNA immunization promoted Th1 immune response and confirmed the previous observations on immunogenicity of LACK and TSA antigens against CL. Furthermore, this study demonstrated that a bivalent vaccine can induce stronger immune responses and protection against infectious challenge with L. major.

Matched MeSH terms: Recombinant Fusion Proteins/immunology; Protozoan Proteins/immunology*
Construction of a recombinant vaccinia virus expressing Babesia gibsoni thrombospondin-related anonymous protein and evaluation of its immunogenicity in mice

Nakamura C, Liu MM, Goo YK, Zhang GH, Jia HL, Kumagai A, et al.

Trop Biomed, 2020 Dec 01;37(4):1029-1037.
PMID: 33612755 DOI: 10.47665/tb.37.4.1029

Previously, we have identified a gene encoding thrombospondin-related anonymous protein of Babesia gibsoni (BgTRAP), and have shown that the antisera raised against recombinant BgTRAP expressed in Escherichia coli inhibited the growth of parasites. In the present study, a recombinant vaccinia virus expressing the BgTRAP (VV/BgTRAP) was constructed. A specific band with a molecular mass of 80 kDa, which is similar to that of native BgTRAP on the merozoites of B. gibsoni, was detected in the supernatant of VV/ BgTRAP-infected RK13 cells. Mice inoculated with VV/BgTRAP produced a specific antiBgTRAP response. The antiserum against VV/BgTRAP showed reactivity against the native BgTRAP on parasites. These results indicated that the recombinant vaccinia virus expressing BgTRAP might be a vaccine candidate against canine B. gibsoni infection.

Matched MeSH terms: Recombinant Proteins/immunology; Protozoan Proteins/immunology*
Immunogenicity of a truncated enterovirus 71 VP1 protein fused to a Newcastle disease virus nucleocapsid protein fragment in mice

Ch'ng WC, Saw WT, Yusoff K, Shafee N

Acta Virol., 2011;55(3):227-33.
PMID: 21978156

Enterovirus 71 (EV71) is one of the viruses that cause hand, foot and mouth disease. Its viral capsid protein 1 (VP1), which contains many neutralization epitopes, is an ideal target for vaccine development. Recently, we reported the induction of a strong immune response in rabbits to a truncated VP1 fragment (Nt-VP1t) displayed on a recombinant Newcastle disease virus (NDV) capsid protein. Protective efficacy of this vaccine, however, can only be tested in mice, since all EV71 animal models thus far were developed in mouse systems. In this study, we evaluated the type of immune responses against the protein developed by adult BALB/c mice. Nt-VP1t protein induced high levels of VP1 IgG antibody production in mice. Purified VP1 antigen stimulated activation, proliferation and differentiation of splenocytes harvested from these mice. They also produced significant levels of IFN-γ, a Th1-related cytokine. Taken together, Nt-VP1t protein is a potent immunogen in adult mice and our findings provide the data needed for testing of its protective efficacy in mouse models of EV71 infections.

Matched MeSH terms: Recombinant Proteins/immunology; Viral Proteins/immunology*; Capsid Proteins/immunology*
Fulltext Progress and challenges toward the development of vaccines against avian infectious bronchitis

Bande F, Arshad SS, Bejo MH, Moeini H, Omar AR

J Immunol Res, 2015;2015:424860.
PMID: 25954763 DOI: 10.1155/2015/424860

Avian infectious bronchitis (IB) is a widely distributed poultry disease that has huge economic impact on poultry industry. The continuous emergence of new IBV genotypes and lack of cross protection among different IBV genotypes have been an important challenge. Although live attenuated IB vaccines remarkably induce potent immune response, the potential risk of reversion to virulence, neutralization by the maternal antibodies, and recombination and mutation events are important concern on their usage. On the other hand, inactivated vaccines induce a weaker immune response and may require multiple dosing and/or the use of adjuvants that probably have potential safety risks and increased economic burdens. Consequently, alternative IB vaccines are widely sought. Recent advances in recombinant DNA technology have resulted in experimental IB vaccines that show promise in antibody and T-cells responses, comparable to live attenuated vaccines. Recombinant DNA vaccines have also been enhanced to target multiple serotypes and their efficacy has been improved using delivery vectors, nanoadjuvants, and in ovo vaccination approaches. Although most recombinant IB DNA vaccines are yet to be licensed, it is expected that these types of vaccines may hold sway as future vaccines for inducing a cross protection against multiple IBV serotypes.

Matched MeSH terms: Viral Envelope Proteins/immunology; Viral Matrix Proteins/immunology; Viral Nonstructural Proteins/immunology
Mimotopes of heat shock proteins of Salmonella enterica serovar Typhi identified from phage-displayed peptide library

Yuen HL, Shamala D, Thong KL

J Infect Dev Ctries, 2008 Aug 30;2(4):313-23.
PMID: 19741295

BACKGROUND: Heat shock proteins (HSPs) are known to be involved in the pathogenesis of Salmonella enterica serovar Typhi (S. Typhi), the causative agent of typhoid fever. The objective of this study was to apply a phage display library to identify mimotopes of two HSPs, HSP90 and DnaK in S. Typhi.
METHODOLOGY: A 12-mer random peptide library expressed on the surface of the filamentous phage, M13, was used to select the mimotopes of two S. Typhi heat shock proteins by biopanning with monoclonal antibodies (mAbs), DnaK and HSP90. The immunogenicity of the selected peptides was determined through binding affinity with polyclonal antibodies from pooled typhoid-confirmed patients' sera and purified HSPs mAb using Western blotting and ELISA.
RESULTS: Five rounds of biopanning resulted in enrichment of phage clones expressing the binding motifs TDxSTRP and FPSHYWLYPPPT, respectively. The selected peptides showed strong immunoreactivity with patients' sera. Thus, monoclonal antibodies against HSP and patient sera can select common mimotopes from the random peptide library.
CONCLUSION: These findings may provide fundamental information for further studies on diagnostic application or vaccine design against this aetiologic agent of typhoid fever.

Matched MeSH terms: HSP90 Heat-Shock Proteins/immunology*
Fulltext Antigenic cell associated dengue 2 virus proteins detected in vitro using dengue fever patients sera

Abubakar S, Azila A, Suzana M, Chang LY

Malays J Pathol, 2002 Jun;24(1):29-36.
PMID: 16329553

At least three major antigenic dengue 2 virus proteins were recognized by pooled dengue fever patients' sera in infected Aedes albopictus (C6/36) mosquito cells. Dengue virus envelope (E), premembrane (PrM) and non-structural protein 1 (NS 1) dimer were detected beginning on day 3 postinfection in both the cell membrane and cytosolic fractions. Using the patients' sera, the presence of antigenic intermediate core protein (C)-PrM and NS1-non-structural protein 2a (NS2a) in the cytoplasmic fraction of dengue 2 virus infected cells was revealed. The presence of a approximately 92 and approximately 84 kDa NS 1 dimer in the membrane (NS 1m) and cytosolic (NS 1c) fractions of C6/36 cells, respectively, was also recognized. Using individual patient's serum, it was further confirmed that all patients' sera contained antibodies that specifically recognized E, NS 1 and PrM present in the dengue 2 virus-infected cell membrane fractions, suggesting that these glycosylated virus proteins were the main antigenic proteins recognized in vivo. Detection of dengue 2 virus C antibody in some patients further suggested that C could be antigenic if presented in vivo.

Matched MeSH terms: Viral Core Proteins/immunology; Viral Envelope Proteins/immunology; Viral Proteins/immunology*; Viral Nonstructural Proteins/immunology
Allergenic proteins of natural rubber latex

Yeang HY, Arif SA, Yusof F, Sunderasan E

Methods, 2002 May;27(1):32-45.
PMID: 12079415 DOI: 10.1016/S1046-2023(02)00049-X

As the living cytoplasm of laticiferous cells, Hevea brasiliensis latex is a rich blend of organic substances that include a mélange of proteins. A small number of these proteins have given rise to the problem of latex allergy. The salient characteristics of H. brasiliensis latex allergens that are recognized by the International Union of Immunological Societies (IUIS) are reviewed. These are the proteins associated with the rubber particles, the cytosolic C-serum proteins and the B-serum proteins that originate mainly from the lutoids. Procedures for the isolation and purification of latex allergens are discussed, from latex collection in the field to various preparative approaches adopted in the laboratory. As interest in recombinant latex allergens increases, there is a need to validate recombinant proteins to ascertain equivalence with their native counterparts when used in immunological studies, diagnostics, and immunotherapy.

Matched MeSH terms: Plant Proteins/immunology
The manufacture of gloves from natural rubber latex

Yip E, Cacioli P

J Allergy Clin Immunol, 2002 Aug;110(2 Suppl):S3-14.
PMID: 12170237 DOI: 10.1067/mai.2002.124499

Gloves that will provide a barrier of protection from infectious organisms are an essential feature of medical practice for the protection of both patients and medical personnel. Natural rubber latex has consistently been the most satisfactory raw material for the manufacture of gloves. Certain latex proteins, carried over into the finished product by inadequate manufacturing processes, may pose a risk of provoking allergic reactions in some patients and medical workers. As with any allergy, the risk depends on the route of exposure and dose. Hence, the method of manufacture, including the means used to coat gloves to make donning easy, can influence the eventual exposure of sensitive people to latex allergens. In this article, we describe the several processes in use and their effects on latex protein content.

Matched MeSH terms: Plant Proteins/immunology

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