Microbial cell disruption methods for efficient release of enzyme L-asparaginase

Autor(es)

Costa-Silva, Tales A. ^[1] ; Flores-Santos, Juan Carlos ^[1] ; Freire, Rominne K. B. ^[1] ; Vitolo, Michele ^[1] ; Pessoa-, Jr., Adalberto ^[1]

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Dept Pharmaceut & Biochem Technol, Sch Pharmaceut Sci, Av Prof Lineu Prestes 580, B 16, BR-05508900 Sao Paulo, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: PREPARATIVE BIOCHEMISTRY & BIOTECHNOLOGY; v. 48, n. 8, p. 707-717, 2018.
• Citações Web of Science: 3

Resumo

The efficacy of a simple laboratory method for cell disruption based on the glass bead stirring, sonication, osmotic shock, freezing and grinding, or use of solvents and detergents was assessed in this study, via measurements of the release of total protein and L-asparaginase activity. Three different microbial sources of L-asparaginase were used: Escherichia coli BL21 (DE3), Leucosporidium muscorum, and Aspergillus terreus (CCT 7693). This study adjusted and identified the best procedure for each kind of microorganism. Sonication and glass bead stirring led to obtaining filamentous fungus cell-free extracts containing high concentrations of soluble proteins and specific activity; however, sonication was the best since it obtained 4.61 +/- 0.12IUmg(-1) after 3min of operation time. Mechanical methods were also the most effective for yeast cell disruption, but sonication was the technique which yielded a higher efficiency releasing 7.3 IUtotal compared to glass bead stirring releasing 2.7 IUtotal at the same operation time. For bacterium, sonication proved to be the best procedure due to getting the highest specific activity (9.01IUmg(-1)) and total enzyme activity (61.7IU). The data presented lead to conclude that the mechanical methods appeared to be the most effective for the disintegration of the all microbial cells studies. This is the first report related to the experimental comparison of L-ASNase extraction procedures from different microorganisms, which can also be used for extracting periplasm located enzymes from other organisms. (AU)

• Processo FAPESP: 13/08617-7 - Produção de L-asparaginase extracelular: da bioprospecção à engenharia de um biofármaco antileucêmico
• Beneficiário: Adalberto Pessoa Junior
• Modalidade de apoio: Auxílio à Pesquisa - Temático
• Processo FAPESP: 13/19584-2 - Produção de L-asparaginase extracelular pelo fungo Arpergillus terreus
• Beneficiário: Tales Alexandre da Costa e Silva
• Modalidade de apoio: Bolsas no Brasil - Pós-Doutorado