PEGylation versus glycosylation: effect on the thermodynamics and thermostability of crisantaspase

Author(s)

Torres-Obreque, Karin ; Kleingesinds, Eduardo Krebs ; Santos, Joao H. P. M. ; Carretero, Gustavo ; Rabelo, Jheniffer ; Converti, Attilio ; Monteiro, Gisele ; Pessoa Jr, Adalberto ; Rangel-Yagui, Carlota O.

• Document type: Journal article
• Source: PREPARATIVE BIOCHEMISTRY & BIOTECHNOLOGY; v. N/A, p. 11-pg., 2023-09-11.

Abstract

Thermostability is an important and desired feature of therapeutic proteins and is critical for the success or failure of protein drugs development. It can be increased by PEGylation-binding of poly(ethylene glycol) moieties-or glycosylation-post-translational modification to add glycans. Here, the thermostability and thermodynamic parameters of native, PEGylated, and glycosylated versions of the antileukemic enzyme crisantaspase were investigated. First-order kinetics was found to describe the irreversible deactivation process. Activation energy of the enzyme-catalyzed reaction (E*) was estimated for native, PEGylated, and glycosylated enzyme (10.2, 14.8, and 18.8 kJ mol(-1) respectively). Half-life decreased progressively with increasing temperature, and lon-ger half-life was observed for PEG-crisantaspase (87.74 min) at 50 degrees C compared to the native form (9.79 min). The activation energy of denaturation of PEG-crisantaspase (307.1 kJ mol(-1)) was higher than for crisantaspase (218.1 kJ mol(-1)) and Glyco-crisantaspase (120.0 kJ mol(-1)), which means that more energy is required to overcome the energy barrier of the unfolding process. According to our results, PEG-crisantaspase is more thermostable than its native form, while Glyco-crisantaspase is more thermosensitive. (AU)

• FAPESP's process: 16/22065-5 - N-terminal pegylation of proteins and purification by aqueous two-phase systems
• Grantee: Carlota de Oliveira Rangel Yagui
• Support Opportunities: Regular Research Grants
• FAPESP's process: 17/20384-9 - Development of downstream process of humanized L-asparaginase and its characterization
• Grantee: Eduardo Krebs Kleingesinds
• Support Opportunities: Scholarships in Brazil - Doctorate
• FAPESP's process: 13/08617-7 - Production of extracellular L-asparaginase: from bioprospecting to the engineering of an antileukemic biopharmaceutical
• Grantee: Adalberto Pessoa Junior
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 18/25994-2 - Development of novel platforms for PEGylation of proteins with therapeutic potential using microfluidics
• Grantee: João Henrique Picado Madalena Santos
• Support Opportunities: Scholarships in Brazil - Post-Doctoral