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Activation and competition of lipoylation of H protein and its hydrolysis in a reaction cascade catalyzed by the multifunctional enzyme lipoate-protein ligase A
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  • Xinyi Zhang,
  • Jinglei Nie,
  • Yuanmin Zheng,
  • Jie Ren,
  • An-Ping Zeng
Xinyi Zhang
Beijing University of Chemical Technology
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Jinglei Nie
Beijing University of Chemical Technology
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Yuanmin Zheng
Beijing University of Chemical Technology
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Jie Ren
Beijing University of Chemical Technology
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An-Ping Zeng
Hamburg University of Technology
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Abstract

Protein lipoylation is essential for the function of many key enzymes, but barely studied kinetically. Here, the two-step reaction cascade of H protein lipoylation catalyzed by the multifunctional enzyme lipoate-protein ligase A (LplA) was quantitatively and differentially studied. We discovered new phenomena and unusual kinetics of the cascade: (1) the speed of the first reaction is faster than the second one by two orders of magnitude, leading to high accumulation of the intermediate Lip-AMP; (2) Lip-AMP is hydrolyzed, but only significantly at the presence of H protein and in competition with the lipoylation; (3) both the lipoylation of H protein and its hydrolysis are enhanced by the apo and lipoylated forms of H protein and a mutant without the lipoylation site. A conceptual mechanistic model is proposed to explain these experimental observations in which conformational change of LplA upon interaction with H protein and competitive nucleophilic attacks play key roles.

Peer review status:ACCEPTED

26 Apr 2020Submitted to Biotechnology and Bioengineering
27 Apr 2020Submission Checks Completed
27 Apr 2020Assigned to Editor
03 May 2020Reviewer(s) Assigned
07 Jun 2020Review(s) Completed, Editorial Evaluation Pending
07 Jun 2020Editorial Decision: Revise Major
14 Jul 20201st Revision Received
14 Jul 2020Submission Checks Completed
14 Jul 2020Assigned to Editor
19 Jul 2020Reviewer(s) Assigned
02 Aug 2020Review(s) Completed, Editorial Evaluation Pending
02 Aug 2020Editorial Decision: Accept