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Structure of a phage-encoded SAM hydrolase enzyme provides insights in substrate binding and catalysis
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Biology.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Structural Biology.
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(English)Manuscript (preprint) (Other academic)
Keywords [en]
SAM hydrolase, phage enzyme
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:uu:diva-366689OAI: oai:DiVA.org:uu-366689DiVA, id: diva2:1265277
Available from: 2018-11-22 Created: 2018-11-22 Last updated: 2020-01-29
In thesis
1. The Importance of Being Promiscuous: Understanding enzyme function, specificity, and evolution through structure
Open this publication in new window or tab >>The Importance of Being Promiscuous: Understanding enzyme function, specificity, and evolution through structure
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Enzymes are known to be amazingly specific and efficient catalysts. However, many enzymes also have so-called promiscuous functions, i.e., they are able to catalyze other reactions than their main one. The promiscuous activities are often low, serendipitous, and under neutral selection but if conditions arise that make them beneficial, they can play an important role in the evolution of new enzymes. In this thesis, I present three studies where we have characterized different enzyme families by structural and biochemical methods. The studies demonstrate the occurrence of enzyme promiscuity and its potential role in evolution and organismal adaptation.

In the first study, I describe the characterization of wild type and mutant HisA enzymes from Salmonella enterica. In the first part of this study, we could clarify the mechanistic cycle of HisA by solving crystal structures that showed different conformations of wild type HisA in complex with its labile substrate ProFAR (N´-[(5´-phosphoribosyl)formimino]-5-aminoimidazole-4-carboxamide ribonucleotide). In the second part of this study, structures of mutant enzymes from a real-time evolution study provided us with an atomic-level description of how HisA had evolved a new function. The HisA mutants had acquired TrpF activity, either in addition to (bifunctional generalists) or instead of (TrpF specialists) their HisA activity. In the second study, I present the crystal structure and demonstrate promiscuous activity of the TrpC enzyme from Pseudomonas aeruginosa. The activity data demonstrates that the enzyme can turn over a substrate that lacks a substituent that was previously considered essential for catalysis. In the third study, I present the structural and functional characterization of SAM (S-Adenosyl methionine) hydrolases from bacteriophages. These enzymes were discovered because of their ability to rescue auxotrophic bacteria by inducing expression of a promiscuous bacterial enzyme.  

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2018. p. 63
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1747
Keywords
Enzyme evolution, enzyme promiscuity, X-ray crystallography.
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-366690 (URN)978-91-513-0512-7 (ISBN)
Public defence
2019-01-11, B41, BMC, Husargatan 3, Uppsala, 13:00 (English)
Opponent
Supervisors
Available from: 2018-12-14 Created: 2018-11-22 Last updated: 2019-01-21

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