Global untargeted serum metabolomic analyses nominate metabolic pathways responsive to loss of expression of the orphan metallo β-lactamase, MBLAC1

Abstract

The C. elegans gene swip-10 encodes an orphan metallo β-lactamase that genetic studies indicate is vital for limiting neuronal excitability and viability. Sequence analysis indicates that the mammalian gene Mblac1 is the likely ortholog of swip-10, with greatest sequence identity localized to the encoded protein's single metallo β-lactamase domain. The substrate for the SWIP-10 protein remains unknown and to date no functional roles have been ascribed to MBLAC1, though we have shown that the protein binds the neuroprotective β-lactam antibiotic, ceftriaxone. To gain insight into the functional role of MBLAC1 in vivo, we used CRISPR/Cas9 methods to disrupt N-terminal coding sequences of the mouse Mblac1 gene, resulting in a complete loss of protein expression in viable, homozygous knockout (KO) animals. Using serum from both WT and KO mice, we performed global, untargeted metabolomic analyses, resolving small molecules via hydrophilic interaction chromatography (HILIC) based ultra-performance liquid chromatography, coupled to mass spectrometry (UPLC-MS/MS). Unsupervised principal component analysis reliably segregated the metabolomes of MBLAC1 KO and WT mice, with 92 features subsequently nominated as significantly different by ANOVA, and for which we made tentative and putative metabolite assignments. Bioinformatic analyses of these molecules nominate validated pathways subserving bile acid biosynthesis and linoleate metabolism, networks known to be responsive to metabolic and oxidative stress. Our findings lead to hypotheses that can guide future targeted studies seeking to identify the substrate for MBLAC1 and how substrate hydrolysis supports the neuroprotective actions of ceftriaxone.

Evaluating a targeted multiple reaction monitoring approach to global untargeted lipidomic analyses of human plasma

“…RAW files were analyzed using Progenesis QI (Non-linear Dynamics, Newcastle, UK) following a previously described process. 27 Briefly, all the data files (both sample and QC) were imported and aligned against a full MS QC pool reference and ions related to the intact molecule representative of each compound. Data were normalized using Progenesis QI for all compounds.…”

“…RAW files were analyzed using Progenesis QI (Non‐linear Dynamics, Newcastle, UK) following a previously described process 27 . Briefly, all the data files (both sample and QC) were imported and aligned against a full MS QC pool reference and ions related to the intact molecule ([M + Na] + , [M + K] + , [M + Li] + , [M + H] + , [M + H − H 2 O] + , [M − H] − , [M – H − H 2 O] − , [M + Cl] − ) were selected for data processing and deconvolution.…”

Effects of uteroplacental insufficiency on growth-restricted rats with altered lung development: A metabolomic analysis

“…A one-way ANOVA was conducted using Progenesis QI to identify significant differences between the IUGR and control groups. The fold change (FC) threshold calculated by Progenesis QI from the combined abundance data was used to further filter significant features, with a FC of ≥ 1.2 considered significant ( 13 – 15 ). Volcano plots were used to represent dysregulated metabolites [log2 [FC] vs. –log10 [ p value]].…”

Genetic Variability in Molecular Pathways Implicated in Alzheimer's Disease: A Comprehensive Review

“…Binding with metals is a major function of MBLAC1, encoding for metallo-β-lactamase domain-containing protein in the brain (Fagerberg et al, 2014 ). It is involved in hydrolysis of different substrates and metabolic intermediates (Gibson et al, 2018 ). Electron transport chain in mitochondria enables proton motor force generation via redox reactions.…”