| 2005 |
PREPL is localized in the cytosol and possesses a catalytic triad (Ser470, Asp556, His601); substitution of these predicted catalytic residues by alanines resulted in loss of reactivity with a serine hydrolase-specific activity-based probe, confirming an intact but unique catalytic machinery. Unlike prolyl oligopeptidase and oligopeptidase B, PREPL activity depends only on the carboxyterminal domain. |
Activity-based probe labeling with catalytic-residue alanine mutagenesis; subcellular fractionation |
American journal of human genetics |
High |
16385448
|
| 2005 |
PREPL A (638-residue splice variant) was purified from E. coli and shown to have secondary structure similar to oligopeptidase B; it reacted with diisopropyl fluorophosphate and slowly hydrolyzed an activated ester substrate, confirming a reactive catalytic serine. However, PREPL A did not cleave peptide substrates with a P1 basic residue (Arg/Lys), indicating negligible canonical peptidase activity and suggesting a non-peptidase biological function. Dimerization was observed and may account for enhanced conformational stability. |
Recombinant protein expression and purification, differential-scanning calorimetry, enzyme activity assays with peptide and activated ester substrates, DFP inhibitor labeling |
Cellular and molecular life sciences : CMLS |
High |
16143824
|
| 2013 |
PREPL (cytoplasmic) was identified as a direct interaction partner of the N-terminal 70 amino acids of AP-1 complex subunit μ1A via yeast two-hybrid screen. PREPL overexpression reduced AP-1 membrane binding, while reduced PREPL expression increased AP-1 membrane binding and impaired AP-1 recycling. PREPL-deficient patient cell lines displayed an expanded trans-Golgi network that was rescued by PREPL re-expression. PREPL colocalizes with residual membrane-bound AP-1, functioning as an AP-1 effector regulating AP-1 membrane association and TGN morphology. |
Yeast two-hybrid screen; AP-1 membrane binding assays; PREPL overexpression and knockdown; colocalization by fluorescence microscopy; patient cell line complementation |
Journal of cell science |
High |
23321636
|
| 2014 |
PREPL deficiency causes a congenital myasthenic syndrome with decreased quantal content of the endplate potential and reduced miniature endplate potential amplitude without acetylcholine receptor deficiency or altered endplate geometry, indicating both pre- and postsynaptic defects. The myasthenia is attributed to abrogated PREPL interaction with adaptor protein 1 (AP-1). No PREPL expression was detected in patient muscle and endplates. |
Electrophysiology (in vitro neuromuscular transmission studies), immunoblot, histochemistry, ultrastructural studies in patient with isolated PREPL deficiency |
Neurology |
Medium |
24610330
|
| 2014 |
Deletion of exon 11 from the mouse Prepl gene (which encodes key catalytic amino acids) leads to loss of PREPL protein and reduced Prepl mRNA. PREPL-null mice exhibit significantly reduced growth (shorter and lighter) and neonatal hypotonia as measured by a righting reflex assay, establishing that PREPL is required for normal growth and neonatal muscle tone in vivo. |
PREPL knockout mouse model (exon 11 deletion); body measurement; righting assay for neonatal hypotonia |
PloS one |
Medium |
24586561
|
| 2011 |
A fluorescence polarization activity-based protein profiling (fluopol-ABPP) screen identified selective small-molecule inhibitors of PREPL serine hydrolase activity that block PREPL activity in cells. One inhibitor (1-isobutyl-3-oxo-3,5,6,7-tetrahydro-2H-cyclopenta[c]pyridine-4-carbonitrile) distributed to the brain after administration to mice, confirming PREPL is a functional serine hydrolase amenable to pharmacological inhibition. |
Fluopol-ABPP high-throughput screen (~300,000 compounds); cell-based activity assay; mouse pharmacokinetics |
Journal of the American Chemical Society |
Medium |
21692504
|
| 2009 |
NRF-2 and YY-1 transcription factors cooperatively bind a 243-bp GC-rich bidirectional minimal promoter within the 405-bp intergenic region between PREPL and C2ORF34, driving expression of both genes in an additive manner. Cis-acting repressive elements within this region also contribute to tissue-specific expression. |
Promoter deletion/reporter assays, transcription factor binding studies (gel shift/ChIP implied), expression analysis across tissues |
BMC molecular biology |
Medium |
19575798
|
| 2024 |
Missense variants in PREPL from CMS22 patients do not impair hydrolase activity but reduce binding to known interactors, and structural analysis indicates the variants affect regions involved in intraprotein or protein-protein interactions. Catalytically inactive PREPL p.Ser559Ala cell lines showed that hydrolytic activity of PREPL is required for normal mitochondrial function but NOT for regulating AP-1-mediated transport in the trans-Golgi network, demonstrating that PREPL has separable enzymatic and non-enzymatic functions. |
Biochemical hydrolase activity assays; structural analysis; protein-protein interaction assays; CRISPR catalytic-dead knock-in (Ser559Ala) cell lines; mitochondrial function assays; AP-1 transport assays |
JCI insight |
High |
39078710
|
| 2025 |
In PREPL KO HEK293T cells and Prepl KO mouse brains, global phospholipid composition was largely unchanged, arguing against a direct lipase role for PREPL in (lyso)phospholipid turnover. However, PREPL KO cells accumulated triacylglycerols (TAGs) and increased lipid droplets. PREPL does not localize to peroxisomes, and peroxisome numbers and protein levels were largely unaffected, but KO cells showed elongated peroxisomes. The TAG accumulation and peroxisome elongation are interpreted as secondary consequences of mitochondrial dysfunction (impaired fatty acid β-oxidation) caused by PREPL loss, not a direct lipase function. |
Unbiased lipidomics in KO mouse brain and CRISPR-Cas9 KO cell lines; lipid droplet imaging; peroxisome morphology and protein level analysis; PREPL localization studies |
bioRxivpreprint |
Medium |
bio_10.1101_2025.10.28.685080
|