دورية أكاديمية
أعرض في EDS

Cell-Free Protein Expression by a Reconstituted Transcription-Translation System Energized by Sugar Catabolism.

التفاصيل البيبلوغرافية
العنوان: Cell-Free Protein Expression by a Reconstituted Transcription-Translation System Energized by Sugar Catabolism.
المؤلفون: Sato G; Department of Biosciences & Informatics, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama 223-8522, Japan., Miyazawa S; Department of Biosciences & Informatics, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama 223-8522, Japan., Doi N; Department of Biosciences & Informatics, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama 223-8522, Japan., Fujiwara K; Department of Biosciences & Informatics, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama 223-8522, Japan.
المصدر: Molecules (Basel, Switzerland) [Molecules] 2024 Jun 21; Vol. 29 (13). Date of Electronic Publication: 2024 Jun 21.
نوع المنشور: Journal Article
اللغة: English
بيانات الدورية: Publisher: MDPI Country of Publication: Switzerland NLM ID: 100964009 Publication Model: Electronic Cited Medium: Internet ISSN: 1420-3049 (Electronic) Linking ISSN: 14203049 NLM ISO Abbreviation: Molecules Subsets: MEDLINE
أسماء مطبوعة: Original Publication: Basel, Switzerland : MDPI, c1995-
مواضيع طبية MeSH: Glycolysis* , Protein Biosynthesis* , Cell-Free System* , Transcription, Genetic*, Glucose/metabolism ; Adenosine Triphosphate/metabolism ; Sugars/metabolism ; Glucokinase/metabolism ; Glucokinase/genetics
مستخلص: Cooperation between catabolism and anabolism is crucial for maintaining homeostasis in living cells. The most fundamental systems for catabolism and anabolism are the glycolysis of sugars and the transcription-translation (TX-TL) of DNA, respectively. Despite their importance in living cells, the in vitro reconstitution of their cooperation through purified factors has not been achieved, which hinders the elucidation of the design principle in living cells. Here, we reconstituted glycolysis using sugars and integrated it with the PURE system, a commercial in vitro TX-TL kit composed of purified factors. By optimizing key parameters, such as glucokinase and initial phosphate concentrations, we determined suitable conditions for their cooperation. The optimized system showed protein synthesis at up to 33% of that of the original PURE system. We observed that ATP consumption in upstream glycolysis inhibits TX-TL and that this inhibition can be alleviated by the co-addition of glycolytic intermediates, such as glyceraldehyde 3-phosphate, with glucose. Moreover, the system developed here simultaneously synthesizes a subset of its own enzymes, that is, glycolytic enzymes, in a single test tube, which is a necessary step toward self-replication. As glycolysis and TX-TL provide building blocks for constructing cells, the integrated system can be a fundamental material for reconstituting living cells from purified factors.
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معلومات مُعتمدة: KAKENHI Grant Number JP 18H04565, JP20H04717, JP22H05432, JP22K19299 for K.F. and JP22J11648, JP22KJ2689 for G.S. Japan Society for the Promotion of Science
فهرسة مساهمة: Keywords: bottom-up synthetic biology; cell-free protein synthesis; glycolysis
المشرفين على المادة: IY9XDZ35W2 (Glucose)
8L70Q75FXE (Adenosine Triphosphate)
0 (Sugars)
EC 2.7.1.2 (Glucokinase)
تواريخ الأحداث: Date Created: 20240713 Date Completed: 20240713 Latest Revision: 20240715
رمز التحديث: 20240715
مُعرف محوري في PubMed: PMC11243612
DOI: 10.3390/molecules29132956
PMID: 38998908
قاعدة البيانات: MEDLINE
الوصف
تدمد:1420-3049
DOI:10.3390/molecules29132956