HIGH LEVEL MEASLES VIRUS N PROTEIN EXPRESSION USING A RECOMBINANT BACULOVIRUS: ASSEMBLY OF NUCLEOCAPSID-LIKE STRUCTURES

Epidemiological situation describing global measles spread is ambiguous. Along with countries succeeded in measles eradication, there are those wherein measles rate remains at quite high level. Because measles is a vaccine-preventable infection, it may then be eradicated solely by ensuring sufficient population coverage with preventive vaccination. The aim of our study was to assess level of measles immunity in medical workers at the Clinics of Samara State Medical University as well as the Samara State Medical University. There were enrolled 1503 subjects (aged 18–79 years), among which all individuals under 55 (77.58%) but not older counterparts provided with medical record on previous measles vaccination or measles infection. Level of serum measles virus-specific IgG antibodies was measured by using ELISA (VektoKor-IgG, JSC Vector-Best, Novosibirsk), with mean concentration ranging in general population within 1.02±0.02 IU/ ml. Positive results were observed in 72.52% of the examined individuals. Average vs. high measles virus-specific IgG level was detected in 52.90% (mean age — 41.4±0.5 years) and 19.62% (mean age — 54.2±0.72 years) of individuals, whereas at level below threshold — in 27.48% of subjects (mean age — 33.25±0.53 years). Thus, in 34.16% of the surveyed vaccinated individuals mostly presented by young subjects contained anti-measles virus-specific antibodies below protective level. Older age groups were shown to increase in average IgG amount with age. Interestingly, age-related measles immunity pattern was observed: percentage of subjects with high vs. low measles virus-specific IgG level increases and decreases, respectively. Taking into consideration a large percentage of subjects previously vaccinated against measles among carriers of low measles immunity, it may be concluded that measles virus-specific IgG antibody level must be monitored in young adulthood to decide of whether subsequent revaccination is necessary.

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Moving the Glycoprotein Gene of Vesicular Stomatitis Virus to Promoter-Proximal Positions Accelerates and Enhances the Protective Immune Response

Journal of Virology ◽

10.1128/jvi.74.17.7895-7902.2000 ◽

2000 ◽

Vol 74 (17) ◽

pp. 7895-7902 ◽

Cited By ~ 49

Author(s):

E. Brian Flanagan ◽

L. Andrew Ball ◽

Gail W. Wertz

Keyword(s):

Immune Response ◽

Protein Expression ◽

Vesicular Stomatitis Virus ◽

Wild Type Virus ◽

Wild Type ◽

Gene Encoding ◽

N Protein ◽

Glycoprotein G ◽

Vesicular Stomatitis ◽

Negative Sense

ABSTRACT Vesicular stomatitis virus (VSV) is the prototype of the Rhabdoviridae and contains nonsegmented negative-sense RNA as its genome. The 11-kb genome encodes five genes in the order 3′-N-P-M-G-L-5′, and transcription is obligatorily sequential from the single 3′ promoter. As a result, genes at promoter-proximal positions are transcribed at higher levels than those at promoter-distal positions. Previous work demonstrated that moving the gene encoding the nucleocapsid protein N to successively more promoter-distal positions resulted in stepwise attenuation of replication and lethality for mice. In the present study we investigated whether moving the gene for the attachment glycoprotein G, which encodes the major neutralizing epitopes, from its fourth position up to first in the gene order would increase G protein expression in cells and alter the immune response in inoculated animals. In addition to moving the G gene alone, we also constructed viruses having both the G and N genes rearranged. This produced three variant viruses having the orders 3′-G-N-P-M-L-5′ (G1N2), 3′-P-M-G-N-L-5′ (G3N4), and 3′-G-P-M-N-L-5′ (G1N4), respectively. These viruses differed from one another and from wild-type virus in their levels of gene expression and replication in cell culture. The viruses also differed in their pathogenesis, immunogenicity, and level of protection of mice against challenge with wild-type VSV. Translocation of the G gene altered the kinetics and level of the antibody response in mice, and simultaneous reduction of N protein expression reduced replication and lethality for animals. These studies demonstrate that gene rearrangement can be exploited to design nonsegmented negative-sense RNA viruses that have characteristics desirable in candidates for live attenuated vaccines.

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High-level extracellular protein expression in Bacillus subtilis by optimizing strong promoters based on the transcriptome of Bacillus subtilis and Bacillus megaterium

Protein Expression and Purification ◽

10.1016/j.pep.2018.06.006 ◽

2018 ◽

Vol 151 ◽

pp. 72-77 ◽

Cited By ~ 6

Author(s):

Xin Liu ◽

Hai Wang ◽

Bin Wang ◽

Li Pan

Keyword(s):

Bacillus Subtilis ◽

Protein Expression ◽

Bacillus Megaterium ◽

Extracellular Protein ◽

High Level

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High-Level Conversion of l-lysine into Cadaverine by Escherichia coli Whole Cell Biocatalyst Expressing Hafnia alvei l-lysine Decarboxylase

Polymers ◽

10.3390/polym11071184 ◽

2019 ◽

Vol 11 (7) ◽

pp. 1184 ◽

Cited By ~ 7

Author(s):

Kim ◽

Baritugo ◽

Oh ◽

Kang ◽

Jung ◽

...

Keyword(s):

Escherichia Coli ◽

Protein Expression ◽

Pyridoxal Phosphate ◽

Lysine Decarboxylase ◽

Hafnia Alvei ◽

Whole Cell ◽

E Coli ◽

Whole Cell Biocatalyst ◽

High Level ◽

Whole Cell Bioconversion

Cadaverine is a C5 diamine monomer used for the production of bio-based polyamide 510. Cadaverine is produced by the decarboxylation of l-lysine using a lysine decarboxylase (LDC). In this study, we developed recombinant Escherichia coli strains for the expression of LDC from Hafnia alvei. The resulting recombinant XBHaLDC strain was used as a whole cell biocatalyst for the high-level bioconversion of l-lysine into cadaverine without the supplementation of isopropyl β-d-1-thiogalactopyranoside (IPTG) for the induction of protein expression and pyridoxal phosphate (PLP), a key cofactor for an LDC reaction. The comparison of results from enzyme characterization of E. coli and H. alvei LDC revealed that H. alvei LDC exhibited greater bioconversion ability than E. coli LDC due to higher levels of protein expression in all cellular fractions and a higher specific activity at 37 °C (1825 U/mg protein > 1003 U/mg protein). The recombinant XBHaLDC and XBEcLDC strains were constructed for the high-level production of cadaverine. Recombinant XBHaLDC produced a 1.3-fold higher titer of cadaverine (6.1 g/L) than the XBEcLDC strain (4.8 g/L) from 10 g/L of l-lysine. Furthermore, XBHaLDC, concentrated to an optical density (OD600) of 50, efficiently produced 136 g/L of cadaverine from 200 g/L of l-lysine (97% molar yield) via an IPTG- and PLP-free whole cell bioconversion reaction. Cadaverine synthesized via a whole cell biocatalyst reaction using XBHaLDC was purified to polymer grade, and purified cadaverine was successfully used for the synthesis of polyamide 510. In conclusion, an IPTG- and PLP-free whole cell bioconversion process of l-lysine into cadaverine, using recombinant XBHaLDC, was successfully utilized for the production of bio-based polyamide 510, which has physical and thermal properties similar to polyamide 510 synthesized from chemical-grade cadaverine.

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Measles Virus Assembly within Membrane Rafts

Journal of Virology ◽

10.1128/jvi.74.21.9911-9915.2000 ◽

2000 ◽

Vol 74 (21) ◽

pp. 9911-9915 ◽

Cited By ~ 118

Author(s):

Séverine Vincent ◽

Denis Gerlier ◽

Serge N. Manié

Keyword(s):

G Protein ◽

Measles Virus ◽

Virus Assembly ◽

Membrane Rafts ◽

Virus Envelope ◽

N Protein ◽

Infected Cells ◽

Vesicular Stomatitis ◽

Triton X ◽

N Proteins

ABSTRACT During measles virus (MV) replication, approximately half of the internal M and N proteins, together with envelope H and F glycoproteins, are selectively enriched in microdomains rich in cholesterol and sphingolipids called membrane rafts. Rafts isolated from MV-infected cells after cold Triton X-100 solubilization and flotation in a sucrose gradient contain all MV components and are infectious. Furthermore, the H and F glycoproteins from released virus are also partly in membrane rafts (S. N. Manié et al., J. Virol. 74:305–311, 2000). When expressed alone, the M but not N protein shows a low partitioning (around 10%) into rafts; this distribution is unchanged when all of the internal proteins, M, N, P, and L, are coexpressed. After infection with MGV, a chimeric MV where both H and F proteins have been replaced by vesicular stomatitis virus G protein, both the M and N proteins were found enriched in membrane rafts, whereas the G protein was not. These data suggest that assembly of internal MV proteins into rafts requires the presence of the MV genome. The F but not H glycoprotein has the intrinsic ability to be localized in rafts. When coexpressed with F, the H glycoprotein is dragged into the rafts. This is not observed following coexpression of either the M or N protein. We propose a model for MV assembly into membrane rafts where the virus envelope and the ribonucleoparticle colocalize and associate.

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Measles Virus Circumvents the Host Interferon Response by Different Actions of the C and V Proteins

Journal of Virology ◽

10.1128/jvi.00108-08 ◽

2008 ◽

Vol 82 (17) ◽

pp. 8296-8306 ◽

Cited By ~ 75

Author(s):

Yuichiro Nakatsu ◽

Makoto Takeda ◽

Shinji Ohno ◽

Yuta Shirogane ◽

Masaharu Iwasaki ◽

...

Keyword(s):

Protein Expression ◽

Measles Virus ◽

Rna Synthesis ◽

Virulent Strain ◽

Viral Rna ◽

Interferon Response ◽

Growth Defect ◽

High Morbidity ◽

C Protein ◽

V Protein

ABSTRACT Measles is an acute febrile infectious disease with high morbidity and mortality. The genome of measles virus (MV), the causative agent, encodes two accessory products, V and C proteins, that play important roles in MV virulence. The V but not the C protein of the IC-B strain (a well-characterized virulent strain of MV) has been shown to block the Jak/Stat signaling pathway and counteract the cellular interferon (IFN) response. We have recently shown that a recombinant IC-B strain that lacks C protein expression replicates poorly in certain cell lines, and its growth defect is related to translational inhibition and strong IFN induction. Here, we show that the V protein of the MV IC-B strain also blocks the IFN induction pathway mediated by the melanoma differentiation-associated gene 5 product, thus actively interfering with the host IFN response at two different steps. On the other hand, the C protein per se possesses no activity to block the IFN induction pathway. Our data indicate that the C protein acts as a regulator of viral RNA synthesis, thereby acting indirectly to suppress IFN induction. Since recombinant MVs with C protein defective in modulating viral RNA synthesis or lacking C protein expression strongly stimulate IFN production, in spite of V protein production, both the C and V proteins must be required for MV to fully circumvent the host IFN response.

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High-level protein expression in scalable CHO transient transfection

Biotechnology and Bioengineering ◽

10.1002/bit.22265 ◽

2009 ◽

Vol 103 (3) ◽

pp. 542-551 ◽

Cited By ~ 72

Author(s):

Jianxin Ye ◽

Vanessa Kober ◽

Melanie Tellers ◽

Zubia Naji ◽

Peter Salmon ◽

...

Keyword(s):

Protein Expression ◽

Transient Transfection ◽

High Level

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Measles virus envelope pseudotyped lentiviral vectors transduce quiescent human HSCs at an efficiency without precedent

Blood Advances ◽

10.1182/bloodadvances.2017007773 ◽

2017 ◽

Vol 1 (23) ◽

pp. 2088-2104 ◽

Cited By ~ 23

Author(s):

Camille Lévy ◽

Fouzia Amirache ◽

Anais Girard-Gagnepain ◽

Cecilia Frecha ◽

Francisco J. Roman-Rodríguez ◽

...

Keyword(s):

Fanconi Anemia ◽

Measles Virus ◽

Virus Envelope ◽

Fanconi Anemia Patient ◽

Key Points ◽

Human Hscs ◽

Cytokine Stimulation ◽

High Level ◽

Very High ◽

Anemia Patient

Key PointsH/F-LVs permit very-high-level transduction of human HSCs, even in the complete absence of cytokine stimulation. H/F-LVs outperform VSV-G-LVs for efficient correction of Fanconi anemia patient cells in unfractionated BM.

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