Tailored high-cell-density fed-batch cultivation and induction strategies for optimized production of a low-immunogenic L-asparaginase mutant in Escherichia coli

IF 3.9 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology Pub Date : 2026-06-01 Epub Date: 2026-02-26 DOI:10.1016/j.enzmictec.2026.110836

Larissa Pereira Brumano , Diego Grumbach , Flaviana da Silva Chaves , Gisele Monteiro , Adalberto Pessoa Jr , Solange I. Mussatto

{"title":"Tailored high-cell-density fed-batch cultivation and induction strategies for optimized production of a low-immunogenic L-asparaginase mutant in Escherichia coli","authors":"Larissa Pereira Brumano , Diego Grumbach , Flaviana da Silva Chaves , Gisele Monteiro , Adalberto Pessoa Jr , Solange I. Mussatto","doi":"10.1016/j.enzmictec.2026.110836","DOIUrl":null,"url":null,"abstract":"<div><div>An integrated upstream process was developed for the high-level production of a low-immunogenic <span>L</span>-asparaginase mutant (EcaII P40S/S206C) in <em>Escherichia coli</em>, combining high-cell-density fed-batch cultivation with controlled induction and post-induction nutrient feeding. Biomass accumulation was achieved by exponential substrate feeding, followed by pH-stat-based nutrient supply during the expression phase. Induction protocols were optimized using biomass-specific IPTG dosing (µmol IPTG/g<sub>cell</sub>), applied either as a pulse or via exponential feeding. This combined strategy resulted in a maximum volumetric enzyme activity of 134,516 U/L and a specific productivity of 3387 U/g<sub>cell</sub>, corresponding to 91-fold and 24-fold increases, respectively, compared with standard batch cultivation. Overall, these results indicate that the coordinated control of feeding strategy and induction conditions substantially improves both volumetric and cell-specific productivity of recombinant <span>L</span>-asparaginase and provides a robust upstream framework for the efficient manufacture of improved enzyme variants.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"197 ","pages":"Article 110836"},"PeriodicalIF":3.9000,"publicationDate":"2026-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Enzyme and Microbial Technology","FirstCategoryId":"5","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0141022926000190","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2026/2/26 0:00:00","PubModel":"Epub","JCR":"Q2","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}

引用次数: 0

Abstract

An integrated upstream process was developed for the high-level production of a low-immunogenic L-asparaginase mutant (EcaII P40S/S206C) in Escherichia coli, combining high-cell-density fed-batch cultivation with controlled induction and post-induction nutrient feeding. Biomass accumulation was achieved by exponential substrate feeding, followed by pH-stat-based nutrient supply during the expression phase. Induction protocols were optimized using biomass-specific IPTG dosing (µmol IPTG/g_cell), applied either as a pulse or via exponential feeding. This combined strategy resulted in a maximum volumetric enzyme activity of 134,516 U/L and a specific productivity of 3387 U/g_cell, corresponding to 91-fold and 24-fold increases, respectively, compared with standard batch cultivation. Overall, these results indicate that the coordinated control of feeding strategy and induction conditions substantially improves both volumetric and cell-specific productivity of recombinant L-asparaginase and provides a robust upstream framework for the efficient manufacture of improved enzyme variants.

查看原文本刊更多论文

量身定制高密度补料分批培养和诱导策略，优化大肠杆菌低免疫原性l-天冬酰胺酶突变体的生产。

为在大肠杆菌中高水平生产低免疫原性l-天冬酰胺酶突变体（EcaII P40S/S206C），开发了一种整合的上游工艺，将高密度投料分批培养与控制诱导和诱导后营养喂养相结合。生物量积累是通过指数底物取食实现的，其次是在表达阶段以ph值为基础的养分供应。采用生物质特异性IPTG剂量（µmol IPTG/gcell）优化诱导方案，采用脉冲或指数进料方式。该组合策略导致最大体积酶活性为134,516 U/L，比产率为3387 U/gcell，分别比标准批量培养提高91倍和24倍。总之，这些结果表明，饲养策略和诱导条件的协调控制大大提高了重组l-天冬酰胺酶的体积和细胞特异性生产力，并为高效制造改进的酶变体提供了一个强大的上游框架。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

求助全文

约1分钟内获得全文求助全文

来源期刊

Enzyme and Microbial Technology 生物-生物工程与应用微生物

CiteScore

7.60

自引率

5.90%

发文量

142

审稿时长

38 days

期刊介绍： Enzyme and Microbial Technology is an international, peer-reviewed journal publishing original research and reviews, of biotechnological significance and novelty, on basic and applied aspects of the science and technology of processes involving the use of enzymes, micro-organisms, animal cells and plant cells. We especially encourage submissions on: Biocatalysis and the use of Directed Evolution in Synthetic Biology and Biotechnology Biotechnological Production of New Bioactive Molecules, Biomaterials, Biopharmaceuticals, and Biofuels New Imaging Techniques and Biosensors, especially as applicable to Healthcare and Systems Biology New Biotechnological Approaches in Genomics, Proteomics and Metabolomics Metabolic Engineering, Biomolecular Engineering and Nanobiotechnology Manuscripts which report isolation, purification, immobilization or utilization of organisms or enzymes which are already well-described in the literature are not suitable for publication in EMT, unless their primary purpose is to report significant new findings or approaches which are of broad biotechnological importance. Similarly, manuscripts which report optimization studies on well-established processes are inappropriate. EMT does not accept papers dealing with mathematical modeling unless they report significant, new experimental data.