This volume of "Methods in Enzymology" series is devoted to the cofactor thiamine, also known as vitamin B1, and its phosphorylated derivatives. The biosynthesis of thiamine is discussed in terms of key enzymes and their regulation, the evolution of metabolic pathways, and its importance in plant and yeast metabolism. The importance of quantifying and distinguishing between phosphorylated and unphosphorylated thiamine in animal and human tissues using chromatographic, enzymatic, and nanopore methods is also discussed. In addition to these methods, thiamine riboswitches are a major focus of this book, covering everything from their identification in plants and the elucidation of their structure to their application in detection.
This book presents state-of-the-art methods and detailed protocols intended for all researchers interested in thiamine metabolism or its detection.
This book presents state-of-the-art methods and detailed protocols intended for all researchers interested in thiamine metabolism or its detection.
Table of Contents
Section 1: Thiamine metabolism and assays1. Continuous directed evolution of thiazole synthase
Kristen Van Gelder, Edmar Ramos de Oliveira-Filho, Ulschan Bathe, Anuran Gayen and J Donato Garcia-Garcia
2. Evolutionary history of thiamine biosynthetic pathways in the domains of life and recent distribution and diversification: a bioinformatic approach
Marie-Anne Van Sluys, Henrique Moura-Dias and Naiara Toledo
3. Generation and characterization of thiamin-biofortified plants
Aymeric Goyer, David Handy and Keonny Cosek
4. Experimental approaches to study thiamine triphosphate and vitamin B1 derivative metabolism in plants
Kai Wang
5. ThiN as a versatile domain of transcriptional repressors and catalytic enzymes of thiamine biosynthesis
Julie A. Maupin-Furlow, Nicholas Cox and Katherine Weber
6. Thiamine deficiency on the oleaginous yeast Yarrowia lipolytica
Cong T. Trinh
7. Assessment of thiamine status in humans
Kyly Whitfield, Elizabeth Frank, Kerry S. Jones and Lucien Bettendorff
8. Assay of thiamine pyrophosphokinase activity
Lucien Bettendorff and Pierre Wins
9. Thiamine quantification in fish tissue samples by assaying transketolase activity
Katie Edwards
10. Engineered nanopore biosensor for thiamine quantification
Florian Leonardus Rudolfus Lucas, Carsten Wloka and Giovanni Maglia
Section 2: Riboswitches
11. Identification and characterisation of thiamine pyrophosphate (TPP) riboswitch in Elaeis guineensis
Zetty Norhana Binti Balia Yusof
12. Scaffold engineering enables high-resolution cryo-EM analysis of thiamine pyrophosphate riboswitch architecture and dynamics
Navtej Toor, Daniel Haack, Madison Maille and Boris Rudolfs
13. Design and applications of chimeric antisense oligonucleotides that bind thiamine pyrophosphate mRNAs as antibacterial agents
Robert Penchovsky
14. Functional analysis of thiamine pyrophosphate-responsive riboswitches using a specialized dual-luciferase reporter gene assay
Matthias Mack and Anna H�benthal
15. Mapping small ligand-RNA interactions using transcriptome-wide sequencing
Willem A. Velema and Daphne van den Homberg
16. Monitoring structural dynamics of nascent bacterial riboswitches responding to TPP within transcriptional complexes
Daniel A. Lafontaine, Mirette Hashem and Carlos Penedo
