دورية أكاديمية
أعرض في EDS

Protein engineering a PhotoRNR chimera based on a unifying evolutionary apparatus among the natural classes of ribonucleotide reductases.

التفاصيل البيبلوغرافية
العنوان: Protein engineering a PhotoRNR chimera based on a unifying evolutionary apparatus among the natural classes of ribonucleotide reductases.
المؤلفون: Song DY; Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA 02138., Stubbe J; Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA 02138., Nocera DG; Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA 02138.
المصدر: Proceedings of the National Academy of Sciences of the United States of America [Proc Natl Acad Sci U S A] 2024 Apr 30; Vol. 121 (18), pp. e2317291121. Date of Electronic Publication: 2024 Apr 22.
نوع المنشور: Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.
اللغة: English
بيانات الدورية: Publisher: National Academy of Sciences Country of Publication: United States NLM ID: 7505876 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1091-6490 (Electronic) Linking ISSN: 00278424 NLM ISO Abbreviation: Proc Natl Acad Sci U S A Subsets: MEDLINE
أسماء مطبوعة: Original Publication: Washington, DC : National Academy of Sciences
مواضيع طبية MeSH: Ribonucleotide Reductases*/metabolism , Ribonucleotide Reductases*/chemistry , Ribonucleotide Reductases*/genetics , Protein Engineering*/methods , Escherichia coli*/genetics , Escherichia coli*/metabolism, Catalytic Domain ; Evolution, Molecular ; Models, Molecular ; Escherichia coli Proteins/metabolism ; Escherichia coli Proteins/genetics ; Escherichia coli Proteins/chemistry
مستخلص: Ribonucleotide reductases (RNRs) are essential enzymes that catalyze the de novo transformation of nucleoside 5'-di(tri)phosphates [ND(T)Ps, where N is A, U, C, or G] to their corresponding deoxynucleotides. Despite the diversity of factors required for function and the low sequence conservation across RNRs, a unifying apparatus consolidating RNR activity is explored. We combine aspects of the protein subunit simplicity of class II RNR with a modified version of Escherichia coli class la photoRNRs that initiate radical chemistry with light to engineer a mimic of a class II enzyme. The design of this RNR involves fusing a truncated form of the active site containing α subunit with the functionally important C-terminal tail of the radical-generating β subunit to render a chimeric RNR. Inspired by a recent cryo-EM structure, a [Re] photooxidant is located adjacent to Y 356 [β], which is an essential component of the radical transport pathway in class I RNRs. Combination of this RNR photochimera with cytidine diphosphate (CDP), adenosine triphosphate (ATP), and light resulted in the generation of Y 356 • along with production of deoxycytidine diphosphate (dCDP) and cytosine. The photoproducts reflect an active site chemistry consistent with both the consensus mechanism of RNR and chemistry observed when RNR is inactivated by mechanism-based inhibitors in the active site. The enzymatic activity of the RNR photochimera in the absence of any β metallocofactor highlights the adaptability of the 10-stranded αβ barrel finger loop to support deoxynucleotide formation and accommodate the design of engineered RNRs.
Competing Interests: Competing interests statement:The authors declare no competing interest.
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معلومات مُعتمدة: R01 GM047274 United States GM NIGMS NIH HHS
فهرسة مساهمة: Keywords: 10-stranded αβ barrel; chimeric enzyme; proton-coupled electron transfer; radical transport; ribonucleotide reductase
المشرفين على المادة: EC 1.17.4.- (Ribonucleotide Reductases)
0 (Escherichia coli Proteins)
تواريخ الأحداث: Date Created: 20240422 Date Completed: 20240429 Latest Revision: 20240505
رمز التحديث: 20240505
مُعرف محوري في PubMed: PMC11067019
DOI: 10.1073/pnas.2317291121
PMID: 38648489
قاعدة البيانات: MEDLINE
الوصف
تدمد:1091-6490
DOI:10.1073/pnas.2317291121