Fan Liu, Xiaoyan Sun, Jingwen Zhou, Jianghua Li, Jian Chen, Guocheng Du, Xinrui Zhao
{"title":"在酿酒酵母中通过增强血红素的输入高效地生物合成活性血红蛋白。","authors":"Fan Liu, Xiaoyan Sun, Jingwen Zhou, Jianghua Li, Jian Chen, Guocheng Du, Xinrui Zhao","doi":"10.1111/febs.17199","DOIUrl":null,"url":null,"abstract":"<p>Hemoglobins, with heme as a cofactor, are functional proteins that have extensive applications in the fields of artificial oxygen carriers and foods. Although <i>Saccharomyces cerevisiae</i> is an ideal host for hemoglobin synthesis, it lacks a suitable transport system to utilize additional heme for active expression of hemoglobins, resulting in the cellular aggregation and degradation of the latter. Here, an effective heme importer, heme-responsive gene 4 (Hrg-4), was selected from six candidates through the comparison of effects on the growth rates of Δ<i>hem1 S. cerevisiae</i> strain and the activities of various hemoglobins when supplemented with 5 mg·L<sup>−1</sup> exogenous heme. Additionally, to counter the instability of plasmid-based expression and the metabolic burden introduced from overexpressing Hrg-4, a series of <i>hrg-4</i> integrated strains were constructed and the best engineered strain with five copies of <i>hrg-4</i> was chosen. We found that this engineered strain was associated with an increased binding rate of heme in monomeric leghemoglobin and multimeric human hemoglobin (76.3% and 16.5%, respectively), as well as an enhanced expression of both hemoglobins (52.8% and 17.0%, respectively). Thus, the engineered strain with improved heme uptake can be used to efficiently synthesize other heme-binding proteins and enzymes in <i>S. cerevisiae</i>.</p>","PeriodicalId":94226,"journal":{"name":"The FEBS journal","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"2024-06-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Efficient biosynthesis of active hemoglobins through enhancing the import of heme in Saccharomyces cerevisiae\",\"authors\":\"Fan Liu, Xiaoyan Sun, Jingwen Zhou, Jianghua Li, Jian Chen, Guocheng Du, Xinrui Zhao\",\"doi\":\"10.1111/febs.17199\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p>Hemoglobins, with heme as a cofactor, are functional proteins that have extensive applications in the fields of artificial oxygen carriers and foods. Although <i>Saccharomyces cerevisiae</i> is an ideal host for hemoglobin synthesis, it lacks a suitable transport system to utilize additional heme for active expression of hemoglobins, resulting in the cellular aggregation and degradation of the latter. Here, an effective heme importer, heme-responsive gene 4 (Hrg-4), was selected from six candidates through the comparison of effects on the growth rates of Δ<i>hem1 S. cerevisiae</i> strain and the activities of various hemoglobins when supplemented with 5 mg·L<sup>−1</sup> exogenous heme. Additionally, to counter the instability of plasmid-based expression and the metabolic burden introduced from overexpressing Hrg-4, a series of <i>hrg-4</i> integrated strains were constructed and the best engineered strain with five copies of <i>hrg-4</i> was chosen. We found that this engineered strain was associated with an increased binding rate of heme in monomeric leghemoglobin and multimeric human hemoglobin (76.3% and 16.5%, respectively), as well as an enhanced expression of both hemoglobins (52.8% and 17.0%, respectively). Thus, the engineered strain with improved heme uptake can be used to efficiently synthesize other heme-binding proteins and enzymes in <i>S. cerevisiae</i>.</p>\",\"PeriodicalId\":94226,\"journal\":{\"name\":\"The FEBS journal\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2024-06-12\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"The FEBS journal\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://onlinelibrary.wiley.com/doi/10.1111/febs.17199\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"The FEBS journal","FirstCategoryId":"1085","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1111/febs.17199","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
Efficient biosynthesis of active hemoglobins through enhancing the import of heme in Saccharomyces cerevisiae
Hemoglobins, with heme as a cofactor, are functional proteins that have extensive applications in the fields of artificial oxygen carriers and foods. Although Saccharomyces cerevisiae is an ideal host for hemoglobin synthesis, it lacks a suitable transport system to utilize additional heme for active expression of hemoglobins, resulting in the cellular aggregation and degradation of the latter. Here, an effective heme importer, heme-responsive gene 4 (Hrg-4), was selected from six candidates through the comparison of effects on the growth rates of Δhem1 S. cerevisiae strain and the activities of various hemoglobins when supplemented with 5 mg·L−1 exogenous heme. Additionally, to counter the instability of plasmid-based expression and the metabolic burden introduced from overexpressing Hrg-4, a series of hrg-4 integrated strains were constructed and the best engineered strain with five copies of hrg-4 was chosen. We found that this engineered strain was associated with an increased binding rate of heme in monomeric leghemoglobin and multimeric human hemoglobin (76.3% and 16.5%, respectively), as well as an enhanced expression of both hemoglobins (52.8% and 17.0%, respectively). Thus, the engineered strain with improved heme uptake can be used to efficiently synthesize other heme-binding proteins and enzymes in S. cerevisiae.