Optimization of the production of a polygalacturonase from Aspergillus kawachii cloned in Saccharomyces cerevisiae in batch and fed-batch cultures

ROJAS N.L.; ORTIZ, G. E.;; CHESINI, M.; BARUQUE, D.J.; CAVALITTO, S. F.

FOOD TECHNOLOGY AND BIOTECHNOLOGY

FACULTY FOOD TECHNOLOGY BIOTECHNOLOGY

Lugar: Zagreb; Año: 2011 vol. 49 p. 316 - 321

1330-9862

Polygalacturonases (PG; EC 3.2.1.15) catalyze the hydrolysis of pectin and/or pectic acid and are useful for industrial applications such as juice clarification and pectin extraction. Growth and heterologous expression of recombinant S. cerevisiae which expresses an acidic PG from Aspergillus kawachii was studied in batch and fed batch cultures. Kinetics and stoichiometric parameters of the recombinant yeast were determined in batch cultures in a synthetic medium. In these cultures, the total biomass concentration, protein concentration, and enzyme activity achieved were 2.2 g/l, 10 mg/l, and 3 U/ml, respectively, to give a productivity of 0.06 U/ml.h. In fed batch cultures, various strategies for galactose feeding were used: a) after a glucose growth phase, the addition of a single pulse of galactose which gave a productivity of 0.19 U/ml.h.; b) after a glucose growth phase, a double pulse of galactose at the same final concentration, resulting in a productivity of 0.21 U/ml.h.; c) a simultaneous feeding of glucose and galactose, yielding a productivity of 1.32 U/ml.h. Based on these results, the simultaneous feeding of glucose and galactose was by far the most suitable strategy for the production of this enzyme. Moreover, some biochemical characteristics of the recombinant enzyme such as a MW of ~60 kDa, an isoelectric point of 3.7 and its ability to hydrolyze polygalacturonic acid at pH 2.5 were determined.