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Protein & Peptide Letters

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ISSN (Print): 0929-8665
ISSN (Online): 1875-5305

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Research Article

Biochemical Study of Fibrinolytic Protease from Euphausia superba Possessing Multifunctional Serine Protease Activity

Author(s): Guo-Ying Qian, Gyutae Lim, Shang-Jun Yin, Jun-Mo Yang, Jinhyuk Lee* and Yong-Doo Park*

Volume 28, Issue 6, 2021

Published on: 12 November, 2020

Page: [651 - 664] Pages: 14

DOI: 10.2174/0929866527666201112123714

Price: $65

Abstract

Background: Fibrinolytic protease from Euphausia superba (EFP) was isolated.

Objective: Biochemical distinctions, regulation of the catalytic function, and the key residues of EFP were investigated.

Methods: The serial inhibition kinetic evaluations coupled with measurements of fluorescence spectra in the presence of 4-(2-aminoethyl) benzene sulfonyl fluoride hydrochloride (AEBSF) was conducted. The computational molecular dynamics (MD) simulations were also applied for a comparative study.

Results: The enzyme behaved as a monomeric protein with a molecular mass of about 28.6 kD with Km BApNA = 0.629 ± 0.02 mM and kcat/Km BApNA = 7.08 s-1/mM. The real-time interval measurements revealed that the inactivation was a first-order reaction, with the kinetic processes shifting from a monophase to a biphase. Measurements of fluorescence spectra showed that serine residue modification by AEBSF directly caused conspicuous changes of the tertiary structures and exposed hydrophobic surfaces. Some osmolytes were applied to find protective roles. These results confirmed that the active region of EFP is more flexible than the overall enzyme molecule and serine, as the key residue, is associated with the regional unfolding of EFP in addition to its catalytic role. The MD simulations were supportive to the kinetics data.

Conclusion: Our study indicated that EFP has an essential serine residue for its catalyst function and associated folding behaviors. Also, the functional role of osmolytes such as proline and glycine that may play a role in defense mechanisms from environmental adaptation in a krill’s body was suggested.

Keywords: Fibrinolytic protease, Euphausia superba, kinetics, unfolding, serine residue, osmolytes, molecular dynamics.

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