Characterization of an immunogenic cellulase secreted by Cryptococcus pathogens

Abstract Members of the C. neoformans/C. gattiii species complex are an important cause of serious humans infections, including meningoencephalitis. We describe here a 45 kDa extracellular cellulase purified from culture supernatants of C. neoformans var. neoformans. The N-terminal sequence obtained from the purified protein was used to isolate a clone containing the full-length coding sequence from a C. neoformans var. neoformans (strain B-3501A) cDNA library. Bioinformatics analysis indicated that this gene is present, with variable homology, in all sequenced genomes of the C. neoformans/C. gattii species complex. The cDNA clone was used to produce a recombinant 45 kDa protein in E. coli that displayed the ability to convert carboxymethyl cellulose and was therefore designated as NG-Case (standing for Neoformans Gattii Cellulase). To explore its potential use as a vaccine candidate, the recombinant protein was used to immunize mice and was found capable of inducing T helper type 1 responses and delayed-type hypersensitivity reactions, but not immune protection against a highly virulent C. neoformans var grubii strain. These data may be useful to better understand the mechanisms underlying the ability C. neoformans/C. gattii to colonize plant habitats and to interact with the human host during infection.

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High-yield production in E. coli and characterization of full-length functional p13II protein from human T-cell leukemia virus type 1

Protein Expression and Purification ◽

10.1016/j.pep.2020.105659 ◽

2020 ◽

Vol 173 ◽

pp. 105659

Author(s):

Elka R. Georgieva ◽

Peter P. Borbat ◽

Christina Fanouraki ◽

Jack H. Freed

Keyword(s):

Leukemia Virus ◽

High Yield ◽

Cell Leukemia Virus Type ◽

E Coli ◽

Yield Production ◽

Human T Cell ◽

T Cell Leukemia Virus ◽

High Yield Production

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In vivo characterization of the activities of novel cyclodipeptide oxidases: new tools for increasing chemical diversity of bioproduced 2,5-diketopiperazines in Escherichia coli

Microbial Cell Factories ◽

10.1186/s12934-020-01432-y ◽

2020 ◽

Vol 19 (1) ◽

Author(s):

Fabien Le Chevalier ◽

Isabelle Correia ◽

Lucrèce Matheron ◽

Morgan Babin ◽

Mireille Moutiez ◽

...

Keyword(s):

Escherichia Coli ◽

Biological Activities ◽

Gene Clusters ◽

Chemical Diversity ◽

E Coli ◽

Chemical Structures ◽

In Vivo Characterization ◽

Culture Supernatants

Abstract Background Cyclodipeptide oxidases (CDOs) are enzymes involved in the biosynthesis of 2,5-diketopiperazines, a class of naturally occurring compounds with a large range of pharmaceutical activities. CDOs belong to cyclodipeptide synthase (CDPS)-dependent pathways, in which they play an early role in the chemical diversification of cyclodipeptides by introducing Cα-Cβ dehydrogenations. Although the activities of more than 100 CDPSs have been determined, the activities of only a few CDOs have been characterized. Furthermore, the assessment of the CDO activities on chemically-synthesized cyclodipeptides has shown these enzymes to be relatively promiscuous, making them interesting tools for cyclodipeptide chemical diversification. The purpose of this study is to provide the first completely microbial toolkit for the efficient bioproduction of a variety of dehydrogenated 2,5-diketopiperazines. Results We mined genomes for CDOs encoded in biosynthetic gene clusters of CDPS-dependent pathways and selected several for characterization. We co-expressed each with their associated CDPS in the pathway using Escherichia coli as a chassis and showed that the cyclodipeptides and the dehydrogenated derivatives were produced in the culture supernatants. We determined the biological activities of the six novel CDOs by solving the chemical structures of the biologically produced dehydrogenated cyclodipeptides. Then, we assessed the six novel CDOs plus two previously characterized CDOs in combinatorial engineering experiments in E. coli. We co-expressed each of the eight CDOs with each of 18 CDPSs selected for the diversity of cyclodipeptides they synthesize. We detected more than 50 dehydrogenated cyclodipeptides and determined the best CDPS/CDO combinations to optimize the production of 23. Conclusions Our study establishes the usefulness of CDPS and CDO for the bioproduction of dehydrogenated cyclodipeptides. It constitutes the first step toward the bioproduction of more complex and diverse 2,5-diketopiperazines.

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Purification of recombinant human N-Acetyltransferase type 1 (NAT1) expressed in E. Coli and characterization of its potential role in folate metabolism

Biochemical Pharmacology ◽

10.1016/0006-2952(95)00087-g ◽

1995 ◽

Vol 49 (12) ◽

pp. 1759-1767 ◽

Cited By ~ 62

Author(s):

A. Ward ◽

M.J. Summers ◽

E. Sim

Keyword(s):

Potential Role ◽

Folate Metabolism ◽

E Coli

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Isolation and Characterization of Monoclonal Antibodies Directed Against Different Epitopes of Type-1-like Fimbriae from a Multifimbriated E. coli Strain

Zentralblatt für Bakteriologie ◽

10.1016/s0934-8840(11)80099-x ◽

1990 ◽

Vol 274 (2) ◽

pp. 155-173 ◽

Cited By ~ 1

Author(s):

Birgitta Biggemann ◽

Thomas Bunse ◽

Albrecht Sachsenweger ◽

Wolfgang Opferkuch

Keyword(s):

Monoclonal Antibodies ◽

Coli Strain ◽

E Coli ◽

Isolation And Characterization

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Cloning and Characterization of a Gene Cluster for Cyclododecanone Oxidation in Rhodococcus ruber SC1

Journal of Bacteriology ◽

10.1128/jb.183.21.6478-6486.2001 ◽

2001 ◽

Vol 183 (21) ◽

pp. 6478-6486 ◽

Cited By ~ 74

Author(s):

Kristy Kostichka ◽

Stuart M. Thomas ◽

Katharine J. Gibson ◽

Vasantha Nagarajan ◽

Qiong Cheng

Keyword(s):

Gene Cluster ◽

Dicarboxylic Acids ◽

Rhodococcus Ruber ◽

Cyclic Ketones ◽

E Coli ◽

Flavin Monooxygenase ◽

Dodecanedioic Acid ◽

Cyclic Compounds

ABSTRACT Biological oxidation of cyclic ketones normally results in formation of the corresponding dicarboxylic acids, which are further metabolized in the cell. Rhodococcus ruber strain SC1 was isolated from an industrial wastewater bioreactor that was able to utilize cyclododecanone as the sole carbon source. A reverse genetic approach was used to isolate a 10-kb gene cluster containing all genes required for oxidative conversion of cyclododecanone to 1,12-dodecanedioic acid (DDDA). The genes required for cyclododecanone oxidation were only marginally similar to the analogous genes for cyclohexanone oxidation. The biochemical function of the enzymes encoded on the 10-kb gene cluster, the flavin monooxygenase, the lactone hydrolase, the alcohol dehydrogenase, and the aldehyde dehydrogenase, was determined in Escherichia coli based on the ability to convert cyclododecanone. Recombinant E. colistrains grown in the presence of cyclododecanone accumulated lauryl lactone, 12-hydroxylauric acid, and/or DDDA depending on the genes cloned. The cyclododecanone monooxygenase is a type 1 Baeyer-Villiger flavin monooxygenase (FAD as cofactor) and exhibited substrate specificity towards long-chain cyclic ketones (C11 to C15), which is different from the specificity of cyclohexanone monooxygenase favoring short-chain cyclic compounds (C5 to C7).

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Mannoprotein from Cryptococcus neoformans Promotes T-Helper Type 1 Anticandidal Responses in Mice

Infection and Immunity ◽

10.1128/iai.70.12.6621-6627.2002 ◽

2002 ◽

Vol 70 (12) ◽

pp. 6621-6627 ◽

Cited By ~ 26

Author(s):

Donatella Pietrella ◽

Rosanna Mazzolla ◽

Patrizia Lupo ◽

Lucia Pitzurra ◽

Maria Jesus Gomez ◽

...

Keyword(s):

T Cells ◽

Cryptococcus Neoformans ◽

Delayed Type Hypersensitivity ◽

Interleukin 12 ◽

Antigenic Determinants ◽

T Helper ◽

Survival Times ◽

Helper Type

ABSTRACT We previously demonstrated that mannoprotein (MP) from Cryptococcus neoformans (CnMP) stimulates interleukin-12 production by human monocytes, thus fostering a T-helper type 1 (Th1) protective anticryptococcal response. In this paper we show that CnMP was also able to induce a Candida albicans-directed protective Th1 response. This was demonstrated for mice immunized with CnMP by induction of a delayed-type hypersensitivity (DTH) reaction to C. albicans MP (CaMP) as well as induction of gamma interferon production by CD4+ and CD8+ splenic T cells stimulated in vitro with CaMP. CnMP-immunized mice were also partially protected from lethal systemic challenge with C. albicans, as shown by prolonged median survival times and decreased fungal burden in the kidney. Much evidence supports the validity of these cross-reactive and functional Th1 responses: (i) a non-cross-reactive C. albicans antigen, such as enolase, did not produce a DTH response to CaMP; (ii) passive adoptive transfer of T cells primed with CnMP induced a DTH reaction; (iii) C. neoformans extract elicited a DTH response to CaMP; and (iv) a monoclonal antibody (7H6) directed against a major and immunodominant T-cell-stimulatory 65-kDa MP (MP65) of C. albicans also recognized discrete 100-kDa constituents of C. neoformans extracts, as well as secretory constituents of the fungus. These results suggest the presence of common Th1 antigenic determinants in the mannoproteic material of C. neoformans and C. albicans epitopes, which should be considered in devising common strategies for immunoprophylactic or immunotherapeutic control of the fungi.

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Histamine H1-receptor antagonists with immunomodulating activities: potential use for modulating T helper type 1 (Th1)/Th2 cytokine imbalance and inflammatory responses in allergic diseases

Clinical & Experimental Immunology ◽

10.1111/j.1365-2249.2009.03958.x ◽

2009 ◽

Vol 157 (1) ◽

pp. 27-34 ◽

Cited By ~ 24

Author(s):

T. Okamoto ◽

S. Iwata ◽

K. Ohnuma ◽

N. H. Dang ◽

C. Morimoto

Keyword(s):

Inflammatory Responses ◽

Allergic Diseases ◽

T Helper ◽

H1 Receptor ◽

Histamine H1 Receptor ◽

Th2 Cytokine ◽

Potential Use ◽

Cytokine Imbalance ◽

Helper Type

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Cell penetrable-mouse forkhead box P3 suppresses type 1 T helper cell-mediated immunity in a murine model of delayed-type hypersensitivity

Experimental and Therapeutic Medicine ◽

10.3892/etm.2017.4020 ◽

2017 ◽

Vol 13 (2) ◽

pp. 421-428

Author(s):

Xia Liu ◽

Jun Wang ◽

Hui Wang ◽

Chen Zhou ◽

Qihong Yu ◽

...

Keyword(s):

Murine Model ◽

T Helper Cell ◽

Delayed Type Hypersensitivity ◽

Helper Cell ◽

Cell Mediated Immunity ◽

T Helper ◽

Forkhead Box P3 ◽

Forkhead Box

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Molecular Characterization of Velogenic Avian Avulavirus Type 1 Isolated from Apparently Healthy Emu Birds: Implications to Viral Maintenance and Spread

Indian Journal of Animal Research ◽

10.18805/ijar.b-4350 ◽

2021 ◽

Author(s):

B. Deepthi ◽

D. Ratnamma ◽

R.N. Ramani Pushpa ◽

Shrikrishna Isloor ◽

B.M. Veeregowda ◽

...

Keyword(s):

Fusion Gene ◽

Mrna Levels ◽

Infected Chicken ◽

Cytokine Mrna ◽

Experimental Inoculation ◽

Highly Virulent ◽

Apparently Healthy

Background: Newcastle disease caused by Avian avulavirus type 1 (AAvV-1) is one of the dreadful diseases affecting poultry and other avian species. Wild birds and several domestic birds are recognized as reservoirs of AAvV-1 and probably contribute to the epidemiology of ND in the domesticated poultry. Hence, efforts have been made to understand the virulence and genetic nature of AAvV-1 isolates obtained from apparently healthy Emu birds.Methods: This study details characterization of a velogenic Emu/5 AAvV-1 isolate obtained from an asymptomatic emu flock. Full- length fusion gene was amplified and subsequent phylogenetic analysis was performed. Experimental inoculation of 3-week old chicken with the isolate resulted in virulent ND. Expression of cytokine mRNA levels in spleen of infected chicken at different time points correlated well with the clinical picture, gross and histopathological lesions.Result: To our knowledge this is the first evidence for the role of apparently healthy emu bird acting as a reservoir of velogenic AAvV-1 of subgenotype XIII 2.2 which proved to be highly virulent to chicken. This study further highlights the role of reservoir birds in AAvV-1 transmission and the need for adopting most realistic strategies in counteracting the disease.

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