Tailored Cofactor Traps for the Detection of Hemithioacetal-Forming Pyridoxal Kinases.
Cast your vote
You can rate an item by clicking the amount of stars they wish to award to this item.
When enough users have cast their vote on this item, the average rating will also be shown.
Your vote was cast
Thank you for your feedback
Thank you for your feedback
MetadataShow full item record
AbstractPyridoxal kinases (PLK) are crucial enzymes for the biosynthesis of pyridoxal phosphate, an important cofactor in a plethora of enzymatic reactions. The evolution of these enzymes resulted in different catalytic designs. In addition to the active site, the importance of a cysteine, embedded within a distant flexible lid region, was recently demonstrated. This cysteine forms a hemithioacetal with the pyridoxal aldehyde and is essential for catalysis. Despite the prevalence of these enzymes in various organisms, no tools were yet available to study the relevance of this lid residue. Here, we introduce pyridoxal probes, each equipped with an electrophilic trapping group in place of the aldehyde to target PLK reactive lid cysteines as a mimic of hemithioacetal formation. The addition of alkyne handles placed at two different positions within the pyridoxal structure facilitates enrichment of PLKs from living cells. Interestingly, depending on the position, the probes displayed a preference for either Gram-positive or Gram-negative PLK enrichment. By applying the cofactor traps, we were able to validate not only previously investigated Staphylococcus aureus and Enterococcus faecalis PLKs but also Escherichia coli and Pseudomonas aeruginosa PLKs, unravelling a crucial role of the lid cysteine for catalysis. Overall, our tailored probes facilitated a reliable readout of lid cysteine containing PLKs, qualifying them as chemical tools for mining further diverse proteomes for this important enzyme class.
CitationACS Chem Biol. 2020 Dec 18;15(12):3227-3234. doi: 10.1021/acschembio.0c00787. Epub 2020 Dec 3.
AffiliationHZI,Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7,38124 Braunschweig, Germany.
PublisherAmerican Society for Chemistry (ACS)
JournalACS chemical biology
The following license files are associated with this item:
- Creative Commons
Except where otherwise noted, this item's license is described as Attribution-NonCommercial-ShareAlike 4.0 International
- A subfamily of bacterial ribokinases utilizes a hemithioacetal for pyridoxal phosphate salvage.
- Authors: Nodwell MB, Koch MF, Alte F, Schneider S, Sieber SA
- Issue date: 2014 Apr 2
- On the mechanism of Escherichia coli pyridoxal kinase inhibition by pyridoxal and pyridoxal 5'-phosphate.
- Authors: di Salvo ML, Nogués I, Parroni A, Tramonti A, Milano T, Pascarella S, Contestabile R
- Issue date: 2015 Sep
- Crystal structure and catalytic mechanism of pyridoxal kinase from Pseudomonas aeruginosa.
- Authors: Kim MI, Hong M
- Issue date: 2016 Sep 9
- Structural and functional studies on Salmonella typhimurium pyridoxal kinase: the first structural evidence for the formation of Schiff base with the substrate.
- Authors: Deka G, Kalyani JN, Jahangir FB, Sabharwal P, Savithri HS, Murthy MRN
- Issue date: 2019 Sep
- Pyridoxal 5'-phosphate is a slow tight binding inhibitor of E. coli pyridoxal kinase.
- Authors: Ghatge MS, Contestabile R, di Salvo ML, Desai JV, Gandhi AK, Camara CM, Florio R, González IN, Parroni A, Schirch V, Safo MK
- Issue date: 2012