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Stabilization of Erwinia carotovora and Rhodospirillum rubrum L-Asparaginases in Complexes with Polycations

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Abstract

The effect of chitosan copolymers and polyethyleneimine (PEI) on the specific activity of L-asparaginases of Erwinia carotovora (EwA) and Rhodospirillum rubrum (RrA) at different pH values, as well as the influence of polycations on the mechanisms of thermal inactivation and trypsinolysis of the enzymes, has been studied. Using circular dichroism spectroscopy, the catalytic activities of polyelectrolyte complexes (PEC) were determined, as well as changes in the structure of native enzymes before and after incubation at elevated temperatures. For most PECs, the pH optimum was shifted by 0.5 units towards more acidic values due to the polycationic nature of the polyelectrolytes. The inactivation of EwA and its complexes was found to be associated predominantly with aggregation, while RrA underwent denaturation without aggregation. A stabilizing effect of complex formation was observed: the EwA aggregation was slowed as a result of the formation of PECs with chitosan and its copolymers. The effect of chitosan in stabilizing EwA upon heating was higher than that of RrA. The PEC of RrA with PEI turned out to be optimal: the first order thermal inactivation constant was decreased by 2.2 times compared to the native enzyme and the inactivation constant during trypsinolysis was reduced two-fold. The chitosan-PEG complexation made it possible to maintain or even increase the high level of catalytic activity of the enzymes with simultaneous enhancement of their thermal stability.

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ACKNOWLEDGMENTS

The work was performed using equipment (FTIR Bruker Tensor 27 spectrometer and Jasco J-815 CD spectrometer) provided in accordance with the development program of Moscow State University.

Funding

The work was supported by the development program of Moscow State University.

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Authors and Affiliations

  1. Department of Chemistry, Moscow State University, 119991, Moscow, Russia

    N. D. Dobryakova & E. V. Kudryashova

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  1. N. D. Dobryakova
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  2. E. V. Kudryashova
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Correspondence to E. V. Kudryashova.

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Translated by I. Gordon

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Abbreviations: CD, circular dichroism; CS, chitosan; EwA, Erwinia carotovora L-asparaginase; PBS, phosphate-buffered saline; PEC, polyelectrolyte complex; PEG, polyethylene glycol; PEI, polyethyleneimine; RrA, Rhodospirillum rubrum L-asparaginase.

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Dobryakova, N.D., Kudryashova, E.V. Stabilization of Erwinia carotovora and Rhodospirillum rubrum L-Asparaginases in Complexes with Polycations. Appl Biochem Microbiol 59, 1183–1191 (2023). https://doi.org/10.1134/S000368382309003X

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