Mouse Spexin: (I) NMR Solution Structure, Docking Models for Receptor Binding, and Histological Expression at Tissue Level.

Matthew K H Wong,Kong Hung Sze,Wendy K W Ko,Tao Huang,Zhao-Xiang Bian,Mulan He,Anderson O L Wong

doi:10.3389/fendo.2021.681646

Matthew K H Wong, Kong Hung Sze + Show 5 more

Open Access

https://doi.org/10.3389/fendo.2021.681646

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Abstract

Spexin (SPX), a highly conserved neuropeptide, is known to have diverse functions and has been implicated/associated with pathological conditions, including obesity, diabetes, anorexia nervosa, and anxiety/mood disorders. Although most of the studies on SPX involved the mouse model, the solution structure of mouse SPX, structural aspects for SPX binding with its receptors GalR2/3, and its cellular expression/distribution in mouse tissues are largely unknown. Using CD and NMR spectroscopies, the solution structure of mouse SPX was shown to be in the form of a helical peptide with a random coil from Asn1 to Pro4 in the N-terminal followed by an α-helix from Gln5 to Gln14 in the C-terminus. The molecular surface of mouse SPX is largely hydrophobic with Lys11 as the only charged residue in the α-helix. Based on the NMR structure obtained, docking models of SPX binding with mouse GalR2 and GalR3 were constructed by homology modeling and MD simulation. The models deduced reveal that the amino acids in SPX, especially Asn1, Leu8, and Leu10, could interact with specific residues in ECL1&2 and TMD2&7 of GalR2 and GalR3 by H-bonding/hydrophobic interactions, which provides the structural evidence to support the idea that the two receptors can act as the cognate receptors for SPX. For tissue distribution of SPX, RT-PCR based on 28 tissues/organs harvested from the mouse demonstrated that SPX was ubiquitously expressed at the tissue level with notable signals detected in the brain, GI tract, liver, gonad, and adrenal gland. Using immunohistochemical staining, protein signals of SPX could be located in the liver, pancreas, white adipose tissue, muscle, stomach, kidney, spleen, gonad, adrenal, and hypothalamo-pituitary axis in a cell type-specific manner. Our results, as a whole, not only can provide the structural information for ligand/receptor interaction for SPX but also establish the anatomical basis for our on-going studies to examine the physiological functions of SPX in the mouse model.

Highlights

Spexin (SPX), called neuropeptide Q, is a recent example of identifying novel peptides using bioinformatic approach prior to their purification/functional studies (1, 2)
The mature peptide of SPX is highly conserved from fish to mammals (4, 5), and recent phylogenetic analysis and comparative synteny reveal that SPX is co-evolved with galanin as a result of whole genome duplication occurred during vertebrate evolution (4, 6)
Since small peptides are known to exist in the form of random coil in solution and start to fold into proper conformation for receptor binding in close proximity to the plasma membrane of target cells, a solvent system with proper hydrophobicity, presumably mimicking the microenvironment close to the cell surface, would be required for the nuclear magnetic resonance (NMR) studies for mouse SPX

Summary

Introduction

Spexin (SPX), called neuropeptide Q, is a recent example of identifying novel peptides using bioinformatic approach prior to their purification/functional studies (1, 2). SPX is encoded by the c12orf[39] gene located in chromosome 12 (3) and confirmed to be a secreted peptide by functional expression in cell lines (e.g., b-TC3/COS-7 cells) (1, 3), probably after protein processing in Golgi and endoplasmic reticulum (3). Functional studies confirm that the galanin receptors GalR2 and GalR3 but not GalR1 can act as the cognate receptors for SPX (6). To a less extent in human studies, SPX was found to be widely expressed at tissue level (7, 8) and involved in diverse functions, including stomach contraction (1), GI tract movement (9), energy balance and weight loss (10), bile acid synthesis (11), appetite control (12, 13), glucose homeostasis (14, 15), lipid metabolism (16, 17), reproduction (18, 19), pain perception (20, 21), stress/anxiety (22, 23), and cardiovascular/renal functions (24). Notable changes in SPX expression/serum level can be associated with pathological conditions/diseases, e.g., in type I/II (7, 26) or gestational diabetes (27), childhood (28, 29) and adult obesity (30), metabolic syndrome (31), cardiovascular disease (32), and anorexia nervosa/other psychiatric disorders (33, 34), which have aroused the interest of using SPX as a new target for drug design with clinical implications (22, 35)

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