The optimized signal peptide and feeding strategy to enhance secretory expression of LegH in Komagataella phaffii

IF 3.7 3区生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY

Process Biochemistry Pub Date : 2025-06-09 DOI:10.1016/j.procbio.2025.06.005

Hongtao Chen , Yueheng Niu , Liuxia Zhang , Yunpeng Wang , Ao Luo , Zhihui Dai , Guocheng Du , Xinrui Zhao

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Abstract

Soy Leghemoglobin (LegH) is a type of heme-binding protein that can confer the meat-like color and a unique flavor to the artificial food. The synthesized LegH in Komagataella Phaffii has been approved by FDA to be used as a food additive in plant-based meat. In this study, K. phaffii X33 strain was used as a host to secrete LegH and a mutated α-Mat signal peptide (V50A) was obtained to increase the secretory efficiency by 150 %. Next, the methanol feeding strategy was optimized and the titer of LegH reached 1.1 g/L in a 5 L fermenter. Based on the previous studies, P1H9-V50A-LegH strain was constructed and the μ-STAT strategy with the supplement of 50 g/L sorbitol was used to further increase the secretory titer of LegH to 4.5 g/L. In the following, the double-copy number of LegH expression cassette was applied and the final titer reached 6.0 g/L under the control of μ-STAT strategy.

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优化信号肽及摄食策略对提高法菲小鹿LegH分泌表达的影响

大豆血红蛋白（LegH）是一种血红素结合蛋白，可以赋予人造食品类似肉类的颜色和独特的风味。在Komagataella Phaffii中合成的LegH已被FDA批准用作植物性肉类的食品添加剂。本研究以K. phaffii X33菌株为宿主分泌LegH，获得突变α-Mat信号肽（V50A），使分泌效率提高150% %。其次，优化甲醇进料策略，在5 L的发酵罐中，LegH滴度达到1.1 g/L。在前人研究的基础上，构建了P1H9-V50A-LegH菌株，采用μ-STAT策略，添加50 g/L山梨醇，进一步提高LegH的分泌滴度至4.5 g/L。随后，在μ-STAT策略控制下，应用双拷贝数的LegH表达盒，最终滴度达到6.0 g/L。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Process Biochemistry 生物-工程：化工

CiteScore

8.30

自引率

4.50%

发文量

374

审稿时长

53 days

期刊介绍： Process Biochemistry is an application-orientated research journal devoted to reporting advances with originality and novelty, in the science and technology of the processes involving bioactive molecules and living organisms. These processes concern the production of useful metabolites or materials, or the removal of toxic compounds using tools and methods of current biology and engineering. Its main areas of interest include novel bioprocesses and enabling technologies (such as nanobiotechnology, tissue engineering, directed evolution, metabolic engineering, systems biology, and synthetic biology) applicable in food (nutraceutical), healthcare (medical, pharmaceutical, cosmetic), energy (biofuels), environmental, and biorefinery industries and their underlying biological and engineering principles.