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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Functional and structural evaluation of the antileukaemic enzymel-asparaginase II expressed at low temperature by differentEscherichia colistrains

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Author(s):
de Moura, Werner Alfinito Feio [1] ; Schultz, Leonardo [1] ; Breyer, Carlos Alexandre [1] ; de Oliveira, Ana Laura Pires [1] ; Tairum, Carlos Abrunhosa [1] ; Fernandes, Gabriella Costa [1] ; Toyama, Marcos Hikari [1] ; Pessoa-Jr, Adalberto ; Monteiro, Gisele [2] ; de Oliveira, Marcos Antonio [1]
Total Authors: 10
Affiliation:
[1] Sao Paulo State Univ UNESP, Inst Biosci, Coastal Campus, BR-11330900 Sao Paulo - Brazil
[2] Pessoa-Jr, Jr., Adalberto, Univ Sao Paulo, Biochem Pharmaceut Technol Dept, Fac Pharmaceut Sci, BR-05508000 Sao Paulo - Brazil
Total Affiliations: 2
Document type: Journal article
Source: Biotechnology Letters; JUL 2020.
Web of Science Citations: 0
Abstract

Acute lymphoblastic leukaemia (ALL) affects lymphoblastic cells and is the most common neoplasm during childhood. Among the pharmaceuticals used in the treatment protocols for ALL, Asparaginase (ASNase) fromEscherichia coli(EcAII) is an essential biodrug. Meanwhile, the use of EcAII in neoplastic treatments causes several side effects, such as immunological reactions, hepatotoxicity, neurotoxicity, depression, and coagulation abnormalities. Commercial EcAII is expressed as a recombinant protein, similar to novel enzymes from different organisms; in fact, EcAII is a tetrameric enzyme with high molecular weight (140 kDa), and its overexpression in recombinant systems often results in bacterial cell death or the production of aggregated or inactive EcAII protein, which is related to the formation of inclusion bodies. On the other hand, several commercial expression strains have been developed to overcome these expression issues, but no studies on a systematic evaluation of theE. colistrains aiming to express recombinant asparaginases have been performed to date. In this study, we evaluated eleven expression strains at a low temperature (16 degrees C) with different characteristics to determine which is the most appropriate for asparaginase expression; recombinant wild-type EcAII (rEcAII) was used as a prototype enzyme and the secondary structure content, oligomeric state, aggregation and specific activity of the enzymes were assessed. Structural analysis suggested that a correctly folded tetrameric rEcAII was obtained using ArcticExpress (DE3), a strain that co-express chaperonins, while all other strains produced poorly folded proteins. Additionally, the enzymatic assays showed high specific activity of proteins expressed by ArcticExpress (DE3) when compared to the other strains used in this work. (AU)

FAPESP's process: 13/08617-7 - Production of extracellular L-asparaginase: from bioprospecting to the engineering of an antileukemic biopharmaceutical
Grantee:Adalberto Pessoa Junior
Support type: Research Projects - Thematic Grants
FAPESP's process: 17/19942-7 - Search for inhibitors of the peroxirredoxin system from pathogens and humans
Grantee:Marcos Antonio de Oliveira
Support type: Regular Research Grants
FAPESP's process: 17/25272-4 - Asparaginase 1 from Saccharomyces Cerevisiae: investigation of the functional and structural effects of the N-terminal domain
Grantee:Gabriella Costa Fernandes
Support type: Scholarships in Brazil - Scientific Initiation
FAPESP's process: 19/04054-4 - Search of inhibitors of typical 2-Cys peroxirredoxin system from eukaryotes: functional and structural evaluation
Grantee:Ana Laura Pires de Oliveira
Support type: Scholarships in Brazil - Master
FAPESP's process: 14/22039-9 - Functional and structural characterization and rational modification of ASPaseM: a new pharmaceutical for the acute lymphoblastic leukemia treatment?
Grantee:Leonardo Schultz da Silva
Support type: Scholarships in Brazil - Doctorate (Direct)
FAPESP's process: 16/19245-1 - Evaluation of bacterial strains to L-asparaginase super expression: an antileukemic enzime from Escherichia coli
Grantee:Werner Alfinito Feio de Moura
Support type: Scholarships in Brazil - Scientific Initiation
FAPESP's process: 17/20291-0 - Characterization of the proinflammatory activity of a new serine protease (cdtsp-2) purified from the total venom of Crotalus durissus terrificus
Grantee:Marcos Hikari Toyama
Support type: Regular Research Grants
FAPESP's process: 11/13500-6 - Investigation of the molecular determinants involved in the interaction with substrates of Tsa1 and Tsa2 of Saccharomyces cerevisiae
Grantee:Carlos Alexandre Breyer
Support type: Scholarships in Brazil - Doctorate