Effect of high pressure on structural modifications and enzymatic activity of a purified X-prolyl dipeptidyl aminopeptidase from Streptococcus thermophilus

M Giannoglou; Z Alexandrakis; Ph Stavros; G Katsaros; P Katapodis; G Nounesis; P Taoukis

doi:10.1016/j.foodchem.2017.12.037

Effect of high pressure on structural modifications and enzymatic activity of a purified X-prolyl dipeptidyl aminopeptidase from Streptococcus thermophilus

Food Chem. 2018 May 15:248:304-311. doi: 10.1016/j.foodchem.2017.12.037. Epub 2017 Dec 11.

Authors

M Giannoglou¹, Z Alexandrakis², Ph Stavros³, G Katsaros⁴, P Katapodis⁵, G Nounesis⁶, P Taoukis⁷

Affiliations

¹ Laboratory of Food Chemistry and Technology, School of Chemical Engineering, National Technical University of Athens, Zografou 15780, Greece. Electronic address: giannoglou@chemeng.ntua.gr.
² Laboratory of Food Chemistry and Technology, School of Chemical Engineering, National Technical University of Athens, Zografou 15780, Greece. Electronic address: zalexand@chemeng.ntua.gr.
³ Biomolecular Physics Laboratory, National Center for Scientific Research "Demokritos", 15310 Aghia Paraskevi, Greece. Electronic address: fkstavrou@yahoo.com.
⁴ Institute of Technology of Agricultural Products, Hellenic Agricultural Organisation-DEMETER, Lykovrissi 14123, Attica, Greece. Electronic address: gkats@chemeng.ntua.gr.
⁵ Laboratory of Biotechnology, Department of Biological Applications & Technology, University of Ioannina 45110 Ioannina, Greece. Electronic address: pkatapo@cc.uoi.gr.
⁶ Biomolecular Physics Laboratory, National Center for Scientific Research "Demokritos", 15310 Aghia Paraskevi, Greece. Electronic address: nounesis@rrp.demokritos.gr.
⁷ Laboratory of Food Chemistry and Technology, School of Chemical Engineering, National Technical University of Athens, Zografou 15780, Greece. Electronic address: taoukis@chemeng.ntua.gr.

PMID: 29329859
DOI: 10.1016/j.foodchem.2017.12.037

Abstract

PepX aminopeptidase from Streptococcus thermophilus ACA DC 0022, used in Greek Feta cheese manufacturing, was purified. PepX comprises two subunits of equal molecular mass estimated, using SDS-PAGE and native-PAGE electrophoresis, to be 86 kDa. The effects of high pressure processing (100-450 MPa, combined with 20-40 °C) on purified PepX activity and structure were studied. Activation of the enzyme was observed after processing at 100-200 MPa and 20-30 °C. More intense processing conditions led to enzyme inactivation. PepX HP-induced conformational changes were also investigated through application of Circular Dichroism spectroscopy (CD). Pressures up to 200 MPa resulted in a structurally molten globule-like state where PepX maintained its secondary structure but the tertiary structure was substantially affected and enzyme activity increased. Both secondary and tertiary structures were affected severely by higher pressures (450 MPa), which reduced enzyme activity.

Keywords: Circular dichroism; HP activation/inactivation; Structural changes; purified PepX aminopeptidase.

MeSH terms

Bacterial Proteins / chemistry*
Bacterial Proteins / isolation & purification
Bacterial Proteins / metabolism*
Cheese
Circular Dichroism
Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / chemistry*
Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / isolation & purification
Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / metabolism*
Electrophoresis, Polyacrylamide Gel
Molecular Weight
Pressure
Protein Conformation
Streptococcus thermophilus / enzymology*
Temperature

Substances

Bacterial Proteins
Dipeptidyl-Peptidases and Tripeptidyl-Peptidases
PepX dipeptidyl aminopeptidase