The peptide hormone apelin is translated as a 77-residue preproprotein, truncated to the 55-residue proapelin and, subsequently, to 13–36-residue bioactive isoforms named apelin-13 to -36. Proapelin is hypothesized to be cleaved to apelin-36 and then to the shorter isoforms. However, neither the mechanism of proapelin processing nor the endoproteases involved have been determined. We show direct cleavage of proapelin to apelin-13 by proprotein convertase subtilisin/kexin 3 (PCSK3, or furin) in vitro, with no production of longer isoforms. Conversely, neither PCSK1 nor PCSK7 has appreciable proapelin cleavage activity. Furthermore, we show that both proapelin and PCSK3 transcript expression levels are increased in adipose tissue with obesity and during adipogenesis, suggesting that PCSK3 is responsible for proapelin processing in adipose tissue.
Membrane proteins are still heavily underrepresented in the protein data bank (PDB) due to multiple bottlenecks. The typical low abundance of membrane proteins in their natural hosts makes it necessary to overexpress these proteins either in heterologous systems or through in vitro translation/cell-free expression. Heterologous expression of proteins, in turn, leads to multiple obstacles due to the unpredictability of compatibility of the target protein for expression in a given host. The highly hydrophobic and/or amphipathic nature of membrane proteins also leads to challenges in producing a homogeneous, stable, and pure sample for structural studies. Circumventing these hurdles has become possible through introduction of novel protein production protocols; efficient protein isolation and sample preparation methods; and, improvement in hardware and software for structural characterization. Combined, these advances have made the past 10-15 years very exciting and eventful for the field of membrane protein structural biology, with an exponential growth in the number of solved membrane protein structures. In this review, we focus on both the advances and diversity of protein production and purification methods that have allowed this growth in structural knowledge of membrane proteins through X-ray crystallography, nuclear magnetic resonance (NMR) spectroscopy, and cryoelectron microscopy (cryo-EM).
Apelin and apela (ELABELA/ELA/Toddler) are two peptide ligands for a class A G-protein-coupled receptor named the apelin receptor (AR/APJ/APLNR). Ligand-AR interactions have been implicated in regulation of the adipoinsular axis, cardiovascular system, and central nervous system alongside pathological processes. Each ligand may be processed into a variety of bioactive isoforms endogenously, with apelin ranging from 13 to 55 amino acids and apela from 11 to 32, typically being cleaved C-terminal to dibasic proprotein convertase cleavage sites. The C-terminal region of the respective precursor protein is retained and is responsible for receptor binding and subsequent activation. Interestingly, both apelin and apela exhibit isoform-dependent variability in potency and efficacy under various physiological and pathological conditions, but most studies focus on a single isoform. Biophysical behavior and structural properties of apelin and apela isoforms show strong correlations with functional studies, with key motifs now well determined for apelin. Unlike its ligands, the AR has been relatively difficult to characterize by biophysical techniques, with most characterization to date being focused on effects of mutagenesis. This situation may improve following a recently reported AR crystal structure, but there are still barriers to overcome in terms of comprehensive biophysical study. In this review, we summarize the three components of the apelinergic system in terms of structure-function correlation, with a particular focus on isoform-dependent properties, underlining the potential for regulation of the system through multiple endogenous ligands and isoforms, isoform-dependent pharmacological properties, and biological membrane-mediated receptor interaction. © 2018 American Physiological Society. Compr Physiol 8:407-450, 2018.
Beyond providing novel insight into the physiology of this system, re-annotation of proapelin to the bioactive apelin-55 isoform adds to the molecular toolkit for dissection of apelin-AR interactions and expands the repertoire of therapeutic targets for the apelinergic system.
Ghrelin is a 28-amino acid (aa) stomach-derived peptide discovered in 1999 as the endogenous ligand for growth hormone secretagogue-receptor (GHS-R). Ghrelin-producing cells constitute a distinct group of endocrine cells dispersed throughout the gastric mucosa and to a lesser extent in the small intestine and the endocrine pancreas. Ghrelin plasma levels rise during fasting and chronic caloric restriction to stimulate food intake and fat storage and to prevent life-threatening falls in blood glucose. Plasma ghrelin levels decrease after a meal is consumed and in conditions of energy surplus (such as obesity). Ghrelin has emerged as a key player in the regulation of appetite and energy homeostasis. Ghrelin achieves these functions through binding the ghrelin receptor GHS-R in appetite-regulating neurons and in peripheral metabolic organs including the endocrine pancreas. Ghrelin levels are negatively correlated with body mass index (BMI) and insulin resistance. In addition, ghrelin secretion is impaired in obesity and insulin resistance. Several studies highlight an important role for ghrelin in glucose homeostasis. Genetic, immunological, and pharmacological blockade of ghrelin signaling resulted in improved glucose tolerance and insulin sensitivity. Furthermore, exogenous ghrelin administration was shown to decrease glucose-induced insulin release and increase glucose level in both humans and rodents. GHS-R was shown to be expressed in pancreatic β-cells and ghrelin suppressed insulin release via a Ca2+-mediated pathway. In this review, we provide a detailed summary of recent advances in the field that focuses on the role of insulin and insulin resistance in the regulation of ghrelin secretion and on the role of ghrelin in glucose-stimulated insulin secretion (GSIS).
Vitronectin (Vn) is a major component of blood that controls many processes central to human biology. It is a drug target and a key factor in cell and tissue engineering applications, but despite long-standing efforts, little is known about the molecular basis for its functions. Here, we define the domain organization of Vn, report the crystal structure of its carboxyl-terminal domain, and show that it harbors the binding site for theYersinia pestisouter membrane protein Ail, which recruits Vn to the bacterial cell surface to evade human host defenses. Vn forms a single four-bladed β/α-propeller that serves as a hub for multiple functions. The structure explains key features of native Vn and provides a blueprint for understanding and targeting this essential human protein.
Apela (also referred to as ELABELA and toddler) is a peptide hormone that activates the apelin receptor (AR or APJ) to regulate cardiovascular system development and function. Here, we report the first biophysical characterization of three apela isoforms, apela-54, -32, and -11, alongside a monomeric C1S-apela-11 mutant, using circular dichroism (CD) spectropolarimetry and nuclear magnetic resonance (NMR) spectroscopy. The behaviour of apela-54 is consistent with a preprotein containing a hydrophobic, N-terminal signal peptide. The potential for apela-membrane binding, leading to membrane catalyzed interactions with AR, was tested comprehensively for apela-32 and -11 in the presence of membrane-mimetic dodecylphosphocholine (DPC), sodium dodecyl sulfate (SDS), and 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-rac-(1-glycerol)] (LPPG) micelles. According to pulsed-field gradient diffusion NMR experiments, apela-32 interacts with all three micelles. Chemical shift perturbations indicate widespread interactions along apela, with DPC and LPPG micelles inducing short segments with α-helical character at distinct regions. Consistent with these data, ps-ns dynamics along the peptide backbone appear decreased in the presence of micelles. Apela-11 and C1S-apela-11, alternatively, interact preferentially with SDS and LPPG micelles, promoting β-turn character observable by CD. Distinct differences in membrane-interaction propensity are therefore apparent both as a function of apela isoform and of detergent headgroup. These results imply the potential for cell membrane involvement in apela-AR recognition and binding, with the implication that membrane catalysis has distinct functional and regulatory roles throughout the apelinergic system.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.