scholarly journals Purification of Recombinant Human Tyrosinase from Insect Larvae Infected with the Baculovirus Vector

2017 ◽  
pp. 6.15.1-6.15.12 ◽  
Author(s):  
Monika B. Dolinska ◽  
Paul T. Wingfield ◽  
Yuri V. Sergeev
Keyword(s):  
Insect Larvae ◽  
Baculovirus Vector ◽  
Human Tyrosinase

scholarly journals The consequences of deglycosylation of recombinant intra-melanosomal domain of human tyrosinase

2017 ◽  
Vol 399 (1) ◽  
pp. 73-77 ◽  
Author(s):  
Monika B. Dolinska ◽  
Yuri V. Sergeev
Keyword(s):  
Oculocutaneous Albinism ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Multiple Site ◽  
Insect Larvae ◽  
Melanin Biosynthesis ◽  
Human Tyrosinase

AbstractTyrosinase, a melanosomal glycoenzyme, catalyzes initial steps of the melanin biosynthesis. While glycosylation was previously studiedin vivo, we present three recombinant mutant variants of human tyrosinase, which were obtained using multiple site-directed mutagenesis, expressed in insect larvae, purified and characterized biochemically. The mutagenesis demonstrated the reduced protein expression and enzymatic activity due to possible loss of protein stability and protein degradation. However, the complete deglycosylation of asparagine residuesin vitro, including the residue in position 371, interrupts tyrosinase function, which is consistent with a melanin loss in oculocutaneous albinism type 1 (OCA1) patients.

Are Human Tyrosinase and Related Proteins Suitable Targets for Melanoma Therapy?

2016 ◽  
Vol 16 (27) ◽  
pp. 3033-3047 ◽  
Author(s):  
Elina Buitrago ◽  
Renaud Hardré ◽  
Romain Haudecoeur ◽  
Hélène Jamet ◽  
Catherine Belle ◽  
...  
Keyword(s):  
Related Proteins ◽  
Human Tyrosinase ◽  
Melanoma Therapy

Altered phosphorylation pattern of simian virus 40 T antigen expressed in insect cells by using a baculovirus vector.

1990 ◽  
Vol 64 (10) ◽  
pp. 4799-4807 ◽  
Author(s):  
A Höss ◽  
I Moarefi ◽  
K H Scheidtmann ◽  
L J Cisek ◽  
J L Corden ◽  
...  
Keyword(s):  
Insect Cells ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Baculovirus Vector ◽  
Phosphorylation Pattern

scholarly journals Platforms for Production of Protein-Based Vaccines: From Classical to Next-Generation Strategies

Biomolecules ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1072
Author(s):  
Raquel Cid ◽  
Jorge Bolívar
Keyword(s):  
Infectious Diseases ◽  
Mammalian Cells ◽  
Vaccine Development ◽  
Low Cost ◽  
Cost Effective ◽  
Insect Larvae ◽  
Subunit Vaccines ◽  
Effective Strategies ◽  
Alternative Systems ◽  
Inactivated Vaccines

To date, vaccination has become one of the most effective strategies to control and reduce infectious diseases, preventing millions of deaths worldwide. The earliest vaccines were developed as live-attenuated or inactivated pathogens, and, although they still represent the most extended human vaccine types, they also face some issues, such as the potential to revert to a pathogenic form of live-attenuated formulations or the weaker immune response associated with inactivated vaccines. Advances in genetic engineering have enabled improvements in vaccine design and strategies, such as recombinant subunit vaccines, have emerged, expanding the number of diseases that can be prevented. Moreover, antigen display systems such as VLPs or those designed by nanotechnology have improved the efficacy of subunit vaccines. Platforms for the production of recombinant vaccines have also evolved from the first hosts, Escherichia coli and Saccharomyces cerevisiae, to insect or mammalian cells. Traditional bacterial and yeast systems have been improved by engineering and new systems based on plants or insect larvae have emerged as alternative, low-cost platforms. Vaccine development is still time-consuming and costly, and alternative systems that can offer cost-effective and faster processes are demanding to address infectious diseases that still do not have a treatment and to face possible future pandemics.

scholarly journals Need for growing non-Bt cotton refugia to overcome Bt resistance problem in targeted larvae of the cotton bollworms, Helicoverpa armigera and Pectinophora gossypiella

2021 ◽  
Vol 31 (1) ◽  
Author(s):  
Muhammad Rafiq Shahid ◽  
Muhammad Farooq ◽  
Muhammad Shakeel ◽  
Misbah Ashraf ◽  
Zia Ullah Zia ◽  
...  
Keyword(s):  
Helicoverpa Armigera ◽  
Larval Mortality ◽  
Pectinophora Gossypiella ◽  
Bt Cotton ◽  
Insect Larvae ◽  
Plant Parts ◽  
Arthropod Species ◽  
Bt Resistance ◽  
American Bollworm ◽  
Helicoverpa Armígera

Abstract Background The effectiveness of Bacillus thuringiensis (Bt) cotton against target arthropod larvae is decreasing day by day. The comparative effect of Bt expression among Bt cotton varieties and different plant parts was observed against the cotton bollworms: Helicoverpa armigera and Pectinophora gossypiella larvae. Results In the present study, larval mortality of H. armigera was higher than P. gossypiella among selected Bt cultivars. Median lethal concentration (LC50) values were 8.91, 13.4, 14.0, and 36.4 for P. gossypiella, while 5.91, 4.04, 2.37, and 8.26 for H. armigera of FH-142, MNH-886, IR-3701, and FH-Lalazar, respectively. These values depicted that P. gossypiella had more Bt resistance problem than H. armigera larvae. The host range of both targeted insect larvae was different from each other due to the polyphagous feeding nature of the larvae of H. armigera that feed on different host plants, but P. gossypiella attacked only cotton with monophagous feeding habit. It was also notable from results that Bt expression in reproductive parts where the attacked pink bollworm was lower than the American bollworm, so the former had the maximum chance of resistance due to repeated exposure to Bt. Conclusions It was concluded that farmers be advised to follow the practice of growing non-Bt as a refuge crop to reduce the problem of Bt resistance in the target arthropod species.

scholarly journals Tyrosinase Nanoparticles: Understanding the Melanogenesis Pathway by Isolating the Products of Tyrosinase Enzymatic Reaction

2021 ◽  
Vol 22 (2) ◽  
pp. 734
Author(s):  
Paul K. Varghese ◽  
Mones Abu-Asab ◽  
Emilios K. Dimitriadis ◽  
Monika B. Dolinska ◽  
George P. Morcos ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Large Scale ◽  
Magnetic Beads ◽  
Enzymatic Reaction ◽  
Oculocutaneous Albinism ◽  
Transmission Electron ◽  
Hill Kinetics ◽  
Rate Limiting ◽  
Human Tyrosinase

Human Tyrosinase (Tyr) is the rate-limiting enzyme of the melanogenesis pathway. Tyr catalyzes the oxidation of the substrate L-DOPA into dopachrome and melanin. Currently, the characterization of dopachrome-related products is difficult due to the absence of a simple way to partition dopachrome from protein fraction. Here, we immobilize catalytically pure recombinant human Tyr domain (residues 19–469) containing 6xHis tag to Ni-loaded magnetic beads (MB). Transmission electron microscopy revealed Tyr-MB were within limits of 168.2 ± 24.4 nm while the dark-brown melanin images showed single and polymerized melanin with a diameter of 121.4 ± 18.1 nm. Using Hill kinetics, we show that Tyr-MB has a catalytic activity similar to that of intact Tyr. The diphenol oxidase reactions of L-DOPA show an increase of dopachrome formation with the number of MB and with temperature. At 50 °C, Tyr-MB shows some residual catalytic activity suggesting that the immobilized Tyr has increased protein stability. In contrast, under 37 °C, the dopachrome product, which is isolated from Tyr-MB particles, shows that dopachrome has an orange-brown color that is different from the color of the mixture of L-DOPA, Tyr, and dopachrome. In the future, Tyr-MB could be used for large-scale productions of dopachrome and melanin-related products and finding a treatment for oculocutaneous albinism-inherited diseases.

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