Báo cáo khoa học: Temperature and salts effects on the peptidase activities of the recombinant metallooligopeptidases neurolysin and thimet oligopeptidase

Temperature and Salts Effects on the Peptidase Activities of the Recombinant Metallooligopeptidases Neurolysin and Thimet Oligopeptidase

Authors: Vitor Oliveira, Reynaldo Gatti, Vanessa Rioli, Emer S. Ferro, Alberto Spisni, Antonio C. M. Camargo, Maria A. Juliano, Luiz Juliano

Field: Biochemistry / Enzymology

Document Summary: This study investigates the hydrolytic activities of recombinant neurolysin and thimet oligopeptidase (TOP) on internally quenched fluorescent peptides. The research explores how variations in temperature and salt concentration influence the cleavage sites and kinetics of these metallooligopeptidases. The findings reveal that both neurolysin and TOP exhibit distinct preferences for cleavage bonds, which are modulated by the position and nature of amino acid substitutions, as well as by environmental conditions like temperature and salt concentration. Notably, the presence of salts, particularly NaCl, KCl, and Na2SO4, significantly activates TOP and neurolysin, while NH4Cl and NaI show more modest effects or can reduce TOP activity at higher concentrations. The study also examines the thermodynamic parameters of activation, suggesting that structural rearrangements within the enzymes, rather than significant changes in overall secondary structure, likely contribute to the observed modulation of their activities.

Detailed Table of Contents:

• Introduction
• Materials and Methods
• Thimet oligopeptidase (TOP)
• Neurolysin
• Peptide synthesis
• Kinetic assays
• Temperature dependence of the hydrolysis reaction rates of the substrates by neurolysisn and TOP
• Dependence of the hydrolysis reaction rates by neurolysisn and TOP on concentration and chemical nature of salts
• Determination of cleaved bonds
• Amino-acid analysis
• Circular dichroism
• Results
• Kinetic parameters for the hydrolysis of IQF peptide series Abz-GGFLRRXQ-EDDnp by TOP and neurolysin
• Temperature dependence of the substrate hydrolysis by TOP and neurolysin
• Influence of the NaCl on TOP and neurolysin activities
• Influence of the chemical nature of salts on the TOP and neurolysin activities
• Circular dichroism spectra of TOP and neurolysin
• Discussion
• Acknowledgements
• References