Rhodospirillum rubrum L-Asparaginase Conjugates with Polyamines of Improved Biocatalytic Properties as a New Promising Drug for the Treatment of Leukemiaстатья
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Аннотация:L-asparaginase Rhodospirillum rubrum (RrA) is an enzyme (amidohydrolases; EC 3.5.1.1) that catalyzes the L-asparagine hydrolysis reaction to form L-aspartic acid. Due to the shortcomings of existing L-asparaginases from Esherichia coli (EcA) and Erwinia chrysanthemi (ErA), RrA may turn out to be a new promising drug for the treatment of leukemia. RrA has a low homology with EcA and ErA, which makes the enzyme potentially less immunogenic. RrA has pronounced antitumor activity on a number of leukemia cells. However, there is a need to improve the biocatalytic properties of the enzyme. So, in this study, the RrA conjugates with polyamines with different molecular architectures were developed to regulate the catalytic properties of the enzyme. Linear polyethyleneimine (PEI), branched polyethyleneimine, modified with polyethylene glycol (PEI-PEG), and spermine (Spm) were used to obtain conjugates with RrA. It was discovered by gel permeation chromatography that Spm allows the most active tetrameric form of RrA to be obtained and stabilized. Molecular docking was used to study the binding of spermine to RrA subunits. The activity of the RrA conjugates with Spm and PEI-PEG was 23–30% higher than the native enzyme. The pH optimum of the conjugates shifted from 9.0 to 8.5. The conjugates had higher stability: Spm and PEI-PEG reduced the inactivation constant (kin) more than two-fold upon incubation at 53 °С. The conjugate RrA-PEI-PEG reduced the accessibility of trypsin to the protein surface and reduced kin by eight times. The modification of RrA with polyamines made it possible to obtain enzyme preparations with improved biocatalytic properties. These conjugates represent interest for further study as potential therapeutic agents.
