The middle lipin domain adopts a membrane-binding dimeric protein fold

Phospholipid synthesis and fat storage as triglycerides are regulated by lipin phosphatidic acid phosphatases (PAPs), whose enzymatic PAP function requires association with cellular membranes. Using hydrogen deuterium exchange mass spectrometry, we find mouse lipin 1 binds membranes through an N-terminal amphipathic helix, the Ig-like domain and HAD phosphatase catalytic core, and a middle lipin (M-Lip) domain that is conserved in mammalian and mammalian-like lipins. Crystal structures of the M-Lip domain reveal a previously unrecognized protein fold that dimerizes. The isolated M-Lip domain binds membranes both in vitro and in cells through conserved basic and hydrophobic residues.

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Fig:  Structure and dynamics of lipin 1: a Domain architecture of mouse lipin 1 drawn to scale. Mammalian lipins conserve three regions/domains: the N-Lip, M-Lip, and C-Lip. The N-Lip and C-Lip are predicted to co-fold to form two domains: an Ig-like and HAD-like domain. The positions of the N-ter amphipathic helix (orange h), nuclear localization signal (NLS), and conserved Trp-motif (purple W) are indicated. b % deuterium incorporation after a 3 s (on ice) deuterium exposure in the absence of liposomes. Each point represents a single peptide, with them being graphed on the x-axis according to their central residue. A heat map above is color coded according to the legend. cRegions of lipin 1 that showed significant decreases in exchange (defined as >4%, >0.4 Da, and a two-tailed student t-test p < 0.01) in the presence of liposomes are colored in blue according to the legend and shown on a model of the N-Lip and C-Lip regions generated by Phyre2. Regions not modeled are shown as dashed lines. Helical wheel diagram (top right) of the putative N-terminal amphipathic helix. Hydrophobic residues, yellow; polar residues, blue; glycine residues, gray. d % deuterium incorporation of selected peptides at various time points (3, 30, 300, and 3000 s) in the absence and presence of liposomes. Data are presented as mean values ± SDs. n = 3 independent experiments. Most SDs are smaller than the size of the point. e The sum of the # of deuterons protected from exchange in the presence of liposomes across all timepoints is shown. Each point represents a single peptide, with them being graphed on the x-axis according to their central residue. Data are presented as mean values ± SDs.n = 3 independent experiments. Source data are provided as a Source data file.

Deletion of the M-Lip domain in lipin 1 reduces PAP activity, membrane association, and oligomerization, alters subcellular localization, diminishes acceleration of adipocyte differentiation, but does not affect transcriptional co-activation. This establishes the M-Lip domain as a dimeric protein fold that binds membranes and is critical for full functionality of mammalian lipins.

Gu, W., Gao, S., Wang, H. et al. The middle lipin domain adopts a membrane-binding dimeric protein fold. Nat Commun 12, 4718 (2021). https://doi.org/10.1038/s41467-021-24929-5

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