Customizing Functionalized Cofactor Mimics to Study the Human Pyridoxal 5′-Phosphate-Binding Proteome

Fux, Anja; Pfanzelt, Martin; Kirsch, Volker C.; Hoegl, Annabelle; Sieber, Stephan A.

doi:10.1016/j.chembiol.2019.08.003

Cited by 15 publications

(21 citation statements)

References 46 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Size-exclusion chromatography (SEC) revealed that the protein independent of the presence of the affinity tag is predominantly dimeric even under reductive conditions excluding the role of disulfide bonds in this process (Figures 1A and S1A). 20 This observation differs ). 21 (D) Tandem MS (MS/MS) spectra identifying the PLP-binding site of PLPBP (K47) with corresponding score and posterior error probability (PEP; false discovery rate, FDR < 0.01).…”

Section: ■ Results and Discussionmentioning

confidence: 83%

Biochemical and Proteomic Studies of Human Pyridoxal 5′-Phosphate-Binding Protein (PLPBP)

Fux

Sieber

2019

ACS Chem. Biol.

Self Cite

View full text Add to dashboard Cite

The pyridoxal 5′-phosphate-binding protein (PLPBP) is an evolutionarily conserved protein linked to pyridoxal 5′-phosphate-binding. Although mutations in PLPBP were shown to cause vitamin B6-dependent epilepsy, its cellular role and function remain elusive. We here report a detailed biochemical investigation of human PLPBP and its epilepsy-causing mutants by evaluating stability, cofactor binding, and oligomerization. In this context, chemical cross-linking combined with mass spectrometry unraveled an unexpected dimeric assembly of PLPBP. Furthermore, the interaction network of PLPBP was elucidated by chemical cross-linking paired with co-immunoprecipitation. A mass spectrometric analysis in a PLPBP knockout cell line resulted in distinct proteomic changes compared to wild type cells, including upregulation of several cytoskeleton- and cell division-associated proteins. Finally, transfection experiments with vitamin B6-dependent epilepsy-causing PLPBP variants indicate a potential role of PLPBP in cell division as well as proper muscle function. Taken together, our studies on the structure and cellular role of human PLPBP enable a better understanding of the physiological and pathological mechanism of this important protein.

show abstract

Section: ■ Results and Discussionmentioning

confidence: 83%

Biochemical and Proteomic Studies of Human Pyridoxal 5′-Phosphate-Binding Protein (PLPBP)

Fux

Sieber

2019

ACS Chem. Biol.

Self Cite

View full text Add to dashboard Cite

show abstract

“…Plasma PLP is generally considered to be an effective diagnostic indicator of vitamin B6 status in humans . Due to its function in many biological metabolic pathways, PLP can be used as a biomarker to highly metabolic cells such as cancer cells, especially to metastatic cells . Therefore, PLP is demonstrated as a target molecule for tumors diagnosis, such as bladder, colon and breast cancers .…”

Section: Methodsmentioning

confidence: 99%

Supramolecular Tandem Assay for Pyridoxal‐5′‐phosphate by the Reporter Pair of Guanidinocalix[5]Arene and Fluorescein

et al. 2019

View full text Add to dashboard Cite

show abstract

“…PLP‐dependent enzymes are evolutionary diverse, which makes its classification via sequence homology challenging. The group of Sieber has designed a pyridoxal analogue containing an alkyne click tag in the 2’‐position which is taken up by cells and phosphorylated to form the PLP analogue 36 (Figure 8), [53–54] which was accepted by PLP‐dependent enzymes, and by reduction with NaBH 4 led to the formation of stable conjugates. With this approach they could access 73 % of the current S. aureus PLPome and identified many more putative PLP‐dependent enzymes [53] .…”

Section: Activity‐based Protein Profiling Of Various Oxidoreductasesmentioning

confidence: 99%

“…With this approach they could access 73 % of the current S. aureus PLPome and identified many more putative PLP‐dependent enzymes [53] . Similarly, they could study the PLPome in human HEK293 cells and used it for target screening of Vitamin B 6 antagonists [54] …”

Section: Activity‐based Protein Profiling Of Various Oxidoreductasesmentioning

confidence: 99%

Activity‐Based Protein Profiling (ABPP) of Oxidoreductases

Fuerst

Breinbauer

2020

ChemBioChem

View full text Add to dashboard Cite

Over the last two decades, activity‐based protein profiling (ABPP) has been established as a tremendously useful proteomic tool for measuring the activity of proteins in their cellular context, annotating the function of uncharacterized proteins, and investigating the target profile of small‐molecule inhibitors. Unlike hydrolases and other enzyme classes, which exhibit a characteristic nucleophilic residue, oxidoreductases have received much less attention in ABPP. In this minireview, the state of the art of ABPP of oxidoreductases is described and the scope and limitations of the existing approaches are discussed. It is noted that several ABPP probes have been described for various oxidases, but none so far for a reductase, which gives rise to opportunities for future research.

show abstract

Customizing Functionalized Cofactor Mimics to Study the Human Pyridoxal 5′-Phosphate-Binding Proteome

Abstract: Highlights d Enrichment of human vitamin B 6-binding proteins with cofactor-derived probes d In situ target screening of vitamin B 6 antagonists d Comparison of human cell lines suggests cell-typedependent cofactor loading states

Cited by 15 publications

References 46 publications

Biochemical and Proteomic Studies of Human Pyridoxal 5′-Phosphate-Binding Protein (PLPBP)

Biochemical and Proteomic Studies of Human Pyridoxal 5′-Phosphate-Binding Protein (PLPBP)

Supramolecular Tandem Assay for Pyridoxal‐5′‐phosphate by the Reporter Pair of Guanidinocalix[5]Arene and Fluorescein

Activity‐Based Protein Profiling (ABPP) of Oxidoreductases

Contact Info

Product

Resources

About