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首页> 外文期刊>Applied Microbiology and Biotechnology >Stabilization of cellular mitochondrial enzyme complex and sialyltransferase activity through supplementation of 30Kc19 protein
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Stabilization of cellular mitochondrial enzyme complex and sialyltransferase activity through supplementation of 30Kc19 protein

机译：通过补充30Kc19蛋白来稳定细胞线粒体酶复合物和唾液酸转移酶活性

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摘要

In previous studies, 30Kc19, a lipoprotein in silkworm hemolymph, enhanced productivity and glycosylation by expression of a 30Kc19 gene or supplementation with a recombinant 30Kc19 protein. Additionally, 30Kc19 exhibited enzyme-stabilizing and cell-penetrating abilities in vitro. In this study, we hypothesized that supplemented 30Kc19 penetrated into the cell and enhanced the stability of the cellular enzyme. We investigated this using in vitro and cellular assessments. The activity of sialyltransferase (ST) and isolated mitochondrial complex I/III was enhanced with 30Kc19 in dose-dependent manner while initial reaction rate was unchanged, suggesting that 30Kc19 enhanced enzyme stability rather than specific activity. For intracellular enzyme activity assessment, ST activity inside erythropoietin (EPO)-producing Chinese hamster ovary (CHO) cells increased more than 25 % and mitochondrial complex II activity in HeLa cells increased more than 50 % with 30Kc19. The increase in intracellular ST activity resulted in an increase in sialic acid content of glycoproteins produced in CHO cells supplemented with 30Kc19. Similarly, enhanced mitochondrial complex activity increased mitochondrial membrane potential and ATP production in HeLa cells with 30Kc19 by over 50 %. Because 30Kc19 stabilized intracellular enzymes for glycosylation and enhanced protein productivity with supplementation in the culture medium, we expect that 30Kc19 can be a valuable tool for effective industrial recombinant protein production.
机译：在以前的研究中，家蚕血淋巴中的脂蛋白30Kc19通过表达30Kc19基因或补充重组30Kc19蛋白来提高生产力和糖基化。另外，30Kc19在体外表现出酶稳定和穿透细胞的能力。在这项研究中，我们假设补充的30Kc19会渗入细胞并增强细胞酶的稳定性。我们使用体外和细胞评估方法对此进行了调查。 30Kc19以剂量依赖的方式增强了唾液酸转移酶（ST）和分离的线粒体复合物I / III的活性，而初始反应速率未改变，这表明30Kc19增强了酶的稳定性，而不是比活性。对于细胞内酶活性评估，使用30Kc19时，促红细胞生成素（EPO）产生的中国仓鼠卵巢（CHO）细胞内的ST活性增加了25％以上，HeLa细胞中的线粒体复合体II活性增加了50％以上。细胞内ST活性的增加导致补充有30Kc19的CHO细胞中产生的糖蛋白的唾液酸含量增加。同样，增强的线粒体复合物活性可将30Kc19的HeLa细胞中的线粒体膜电位和ATP产量提高50％以上。由于30Kc19稳定了细胞内酶的糖基化作用，并在培养基中补充了蛋白质，从而提高了生产力，因此我们希望30Kc19可以成为有效的工业重组蛋白生产的有价值的工具。

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