Freitas Marcela, Souza Paula, Homem-de-Mello Mauricio, Fonseca-Bazzo Yris M, Silveira Damaris, Ferreira Filho Edivaldo X, Pessoa Junior Adalberto, Sarker Dipak, Timson David, Inácio João, Magalhães Pérola O
Health Sciences School, University of Brasilia, Brasilia 70910-900, Brazil.
Institute of Biological Sciences, University of Brasilia, Brasilia 70910-900, Brazil.
Pharmaceuticals (Basel). 2022 Jun 14;15(6):746. doi: 10.3390/ph15060746.
L-asparaginase is an important enzyme in the pharmaceutical field used as treatment for acute lymphoblastic leukemia due to its ability to hydrolyze L-asparagine, an essential amino acid synthesized by normal cells, but not by neoplastic cells. Adverse effects of L-asparaginase formulations are associated with its glutaminase activity and bacterial origin; therefore, it is important to find new sources of L-asparaginase produced by eukaryotic microorganisms with low glutaminase activity. This work aimed to identify the L-asparaginase gene sequence from , a filamentous fungus isolated from the Brazilian Savanna (Cerrado) soil with low glutaminase activity, and to biosynthesize higher yields of this enzyme in the yeast . The L-asparaginase gene sequence of was identified by homology to L-asparaginases from species of of the section : and . Partial L-asparaginase from , lacking the periplasmic signaling sequence, was cloned, and expressed intracellularly with highest enzymatic activity achieved by a MUT clone cultured in BMM expression medium; a value 5-fold greater than that obtained by native L-asparaginase in cells. To the best of our knowledge, this is the first literature report of the heterologous production of an L-asparaginase from a filamentous fungus by a yeast.
L-天冬酰胺酶是制药领域中的一种重要酶，因其能够水解L-天冬酰胺（一种由正常细胞而非肿瘤细胞合成的必需氨基酸）而被用作急性淋巴细胞白血病的治疗药物。L-天冬酰胺酶制剂的不良反应与其谷氨酰胺酶活性和细菌来源有关；因此，寻找由谷氨酰胺酶活性低的真核微生物产生的L-天冬酰胺酶的新来源很重要。这项工作旨在从一种从巴西稀树草原（塞拉多）土壤中分离出的谷氨酰胺酶活性低的丝状真菌中鉴定L-天冬酰胺酶基因序列，并在酵母中生物合成更高产量的这种酶。通过与来自节的物种的L-天冬酰胺酶同源性鉴定了的L-天冬酰胺酶基因序列：和。克隆了来自缺少周质信号序列的部分L-天冬酰胺酶，并在胞内表达，通过在BMM表达培养基中培养的MUT克隆实现了最高酶活性；该值比细胞中天然L-天冬酰胺酶获得的值高5倍。据我们所知，这是酵母异源生产丝状真菌L-天冬酰胺酶的首次文献报道。
