Biochemistry & Analytical Biochemistry

Biochemical properties of the deglycosilated recombinant invertase from Candida guilliermondii MpIIIa expressed on Pichia pastoris

Global Congress on Biochemistry, Glycomics & Amino Acids

December 08-09, 2016 San Antonio, USA

Maria Eugenia Hidalgo Lara, Plascencia-Espinosa M A and Santiago-Hernandez J A

CIBA, Mexico
CINVESTAV, Mexico

Posters & Accepted Abstracts: Biochem Anal Biochem

Abstract:

Enzymes are highly effective biological catalysts with great industrial importance because of its versatility and action specificity. It has been reported that glycosylation plays a key role in the biochemical properties of enzymes. The native invertase INV3a-N from Candida guilliermondii MpIIIa is a glycoprotein with a carbohydrate content of ~74%, with optimal activity at pH 5.0 and 65�?°C. In order to improve our knowledge on the crucial role of glycosylation on the biochemical properties of enzymes, the gene inv3a encoding the invertase INV3a-N from C. guilliermondii MpIIIa was expressed in Pichia pastoris X-33, an expression system well known by its low glycosylation of recombinant enzymes. The inv3a gene presented an ORF of 1545 nucleotides encoding a polypeptide of 514 amino acid residues, with a theoretical MW of 58 kDa. The inv3a gene was extracellularly and constitutively expressed in P. pastoris. The recombinant enzyme, named INV3a-R, presented an estimated MW of 66 kDa with a carbohydrate content of ~4-5%, and optimal activity at pH 6.0 and 50�?°C. Interestingly, INV3a-R displays a dramatic diminution in thermal stability at 50�?°C (98.75%), substrate inhibition (50%), Vmax (70%), optimal temperature (23%) and substrate affinity (900%) compared to those values observed for INV3a-N. However, despite being a thermolabile enzyme, INV3a-R is active at pH values up to 6.0, which represents an interesting feature for application in industrial confectionery substrates, whose pH is near 7.0. Findings here confirm the crucial role of glycosylation in the biochemical properties of the invertase INV3a-N from C. guilliermondii MpIIIa.

Biography :

Email: ehidalgo@cinvestav.mx