Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
8328579 | International Journal of Biological Macromolecules | 2018 | 35 Pages |
Abstract
This work reports for the first time the secretory expression of the small laccase (SLAC) from Streptomyces coelicolor A3(2) in Pichia pastoris. Using an AOX1 promoter and α factor as a secretion signal, the recombinant P. pastoris harbouring the laccase gene (rSLAC) produced high titres of extracellular laccase (500â¯Â±â¯10â¯U/l), which were further increased seven fold by pre-incubation at 80â¯Â°C for 30â¯min. The enzyme (â¼38â¯kDa) had an optimum activity at 80â¯Â°C, but optimum pH varied with substrate used. Km values for ABTS, SGZ and 2,6-DMP were 142.85â¯Î¼M, 10â¯Î¼M and 54.55â¯Î¼M and the corresponding kcat values were 60.6â¯sâ1, 25.36â¯sâ1 and 27.84â¯sâ1, respectively. The t1/2 values of the rSLAC at 60â¯Â°C, 70â¯Â°C, 80â¯Â°C were 60â¯h, 32â¯h and 10â¯h, respectively. The enzyme deactivation energy (Ed) was 117.275â¯kJ/mol while ÎG, ÎH and ÎS for thermal inactivation of the rSLAC were all positive. The rSLAC decolourised more than 90% of Brilliant Blue G and Trypan Blue dye in 6â¯h without the addition of a mediator. High titres of SLAC expressed in P. pastoris enhance its potential for various industrial applications.
Keywords
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Authors
Deepti Yadav, Bibhuti Ranjan, Nokuthula Mchunu, Marilize Le Roes-Hill, Tukayi Kudanga,