Peptide self-aggregation and peptide complementarity as bases for the evolution of peptide receptors: a review

This paper reviews the three major theories of peptide receptor evolution: (1) Dwyer's theory that peptide receptors evolved from self-aggregating peptides; (2) Root-Bernstein's theory that peptide receptors evolved from functionally and structurally complementary peptides; and (3) Blalock's theory that receptors evolved from hydropathically complementary sequences encoded in the antisense strand of the DNA encoding each peptide. The evidence to date suggests that the co-yevolution of peptides and their receptors is strongly constrained by one or more of these physicochemically based mechanisms, which argues against a random or frozen accident' model. The data also suggest that structure and function are integrally related from the earliest steps of receptor-ligand evolution so that peptide functionality is non-random and highly conserved in its origin. The result is a molecular paleontology' that reveals the evolutionary constraints that shaped the interaction of structure and function.     

Design of a long acting peptide functioning as both a glucagon-like peptide-1 receptor agonist and a glucagon receptor antagonist

The closely related peptides glucagon-like peptide (GLP-1) and glucagon have opposing effects on blood glucose. GLP-1 induces glucose-dependent insulin secretion in the pancreas, whereas glucagon stimulates gluconeogenesis and glycogenolysis in the liver. The identification of a hybrid peptide acting as both a GLP-1 agonist and a glucagon antagonist would provide a novel approach for the treatment of type 2 diabetes. Toward this end a series of hybrid peptides made up of glucagon and either GLP-1 or exendin-4, a GLP-1 agonist, was engineered. Several peptides that bind to both the GLP-1 and glucagon receptors were identified. The presence of glucagon sequence at the N terminus removed the dipeptidylpeptidase IV cleavage site and increased plasma stability compared with GLP-1. Targeted mutations were incorporated into the optimal dual-receptor binding peptide to identify a peptide with the highly novel property of functioning as both a GLP-1 receptor agonist and a glucagon receptor antagonist. To overcome the short half-life of this mutant peptide in vivo, while retaining dual GLP-1 agonist and glucagon antagonist activities, site-specific attachment of long chained polyethylene glycol (PEGylation) was pursued. PEGylation at the C terminus retained the in vitro activities of the peptide while dramatically prolonging the duration of action in vivo. Thus, we have generated a novel dual-acting peptide with potential for development as a therapeutic for type 2 diabetes.     

Autoreactive T‐cell receptor (Vβ/D/Jβ) sequences in diabetes are homologous to insulin,glucagon, the insulin receptor,and the glucagon receptor

The hypervariable (Vβ/D/Jβ) regions of T‐cell receptors (TCR) have been sequenced in a variety of autoimmune diseases by various investigators. An analysis of some of these sequences shows that TCR from both human diabetics and NOD mice mimic insulin, glucagon, the insulin receptor, and the glucagon receptor. Such similarities are not found in the TCR produced in other human autoimmune diseases. These data may explain how insulin, glucagon, and their receptors are targets of autoimmunity in diabetes and also suggest that TCR mimicking insulin and its receptor may be targets of anti‐insulin autoantibodies. Such intra‐systemic mimicry of self‐proteins also raises complex questions about how “self” and “nonself” are regulated during TCR production, especially in light of the complementarity of insulin for its receptor and glucagon for its receptor. The data presented here suggest that some TCR may be complementary to other TCR in autoimmune diseases, a possibility that is experimentally testable. Such complementarity, if it exists, could either serve to down‐regulate the clones bearing such TCR or, alternatively, trigger an intra‐immune system civil war between them. Copyright © 2008 John Wiley & Sons, Ltd.     

A qualitative comparison of canine plasma gastroenteropancreatic hormone responses to bombesin and the porcine gastrin-releasing peptide (GRP)

The effect on plasma gastroenteropancreatic hormone levels on infusing the porcine gastrin-releasing peptide and bombesin into dogs demonstrated no qualitative difference in the spectrum of activity of the two peptides. Sustained elevations in plasma immunoreactive gastrin, pancreatic polypeptide, enteroglucagon, gastric inhibitory polypeptide, pancreatic glucagon and transient elevations in plasma insulin were seen during infusions of both peptides. The similar spectrum of activities and the structural homology between the two peptides suggests that the porcine gastrin releasing peptide is the porcine counterpart of the amphibian peptide bombesin.     

Vasopressin antisense peptide interactions with the V1 receptor

The molecular recognition hypothesis, that peptide ligands and their receptor binding sites are encoded by complementary nucleotide sequences, was tested for arginine vasopressin (AVP) and its V1 receptor. Binding of [125I] [d(CH2)5,Sar7]AVP (a selective V1 vasopressin antagonist radioligand) or [3H]AVP to rat liver plasma membranes was inhibited by peptides known to bind to V1 receptors but not by the AVP complementary peptide (Ser-Ser-Trp-Ala-Val-Leu-Glu-Val-Ala) (PVA). Rabbit anti-PVA antibodies were nonimmunoreactive with any protein in rat liver membranes or in a partially purified preparation from rat liver containing reconstitutable vasopressin binding activity. Furthermore, there was no suppression of the AVP pressor effect by PVA in vivo using a rat blood pressure bioassay. These findings do not support the hypothesis that the V1 receptor binding site is encoded by the antisense DNA strand to AVP.     

On antisense peptides: the parathyroid hormone as an experimental example and a critical theoretical view

We followed an approach which predicts that translation of two complementary RNA strands into protein generates pairs of "antisense" peptides which bind each other with specific and high affinity (Bost et al. Proc. Natl. Acad. Sci. (1985) 82, 1372). We used human parathormone as an experimental example, and we analysed by computer homologies between antisense peptide sequences and their published receptor sequences. We conclude that there is no experimental indication that parathormone binds to a synthetic peptide, the sequence of which was derived from the antisense RNA sequence. Based on homology scores and antigenicity indexes (Hopp) the analysis shows that the peptide ligand itself, or a random artificial peptide, are as good candidates as the antisense peptide in producing antibodies, presumably recognizing the receptor. We therefore question the general applicability of this approach.     

Targeting the insulin receptor with hormone and peptide dimers

Insulin is a key hormone involved in the regulation of overall energetic homeostasis of the organism. The dimeric character of the receptor for insulin evokes ideas about its activation or inhibition with peptide dimers that could either trigger or block the structural transition of the insulin receptor, leading to its activation. Herewith, we present the chemical engineering and biological characterization of several series of insulin dimers or dimers of specific peptides that should be able to bind receptors for insulin or insulin growth factor 1. The hormones or peptides in the dimers were interconnected with different linkers, consisting of triazole moieties and 3, 6, 8, 11, or 23 polyethylene glycol units. The prepared dimers were weaker in binding to insulin receptors than human insulin. However, some of the insulin dimers showed preferential binding specificity toward the isoform A of the insulin receptor, and the insulin dimers also stimulated the insulin receptor more strongly than would be consistent with their binding affinities. Our results suggest that designing insulin dimers may be a promising strategy for modulating the ability of the hormone to activate the receptor or to alter its specificity toward insulin receptor isoforms.     

Receptors for cholecystokinin and gastrin peptides display specific binding properties and are structurally different in guinea-pig and dog pancreas

In the light of the strong potency of gastrin-related peptides on pancreatic exocrine secretion in dog, we analyzed the binding properties of peptides related to cholecystokinin (CCK) and gastrin on dog pancreatic acini compared to guinea-pig acini. Moreover, we determined apparent molecular masses of photoaffinity labelled CCK/gastrin receptors in the two models. Using the CCK radioligand, receptor selectivity towards CCK/gastrin agonists and antagonists was found to be lower in dog acini than in guinea-pig acini. Performing the binding with CCK and gastrin radioligands in combination with N2,O2'-dibutyryl-guanosine 3',5'-monophosphate, revealed that in dog acini there exist two different sub-classes of CCK/gastrin receptors having high and low selectivity, the latter ones being able to bind gastrin with high affinity (Kd = 2.1 nM). SDS-PAGE analysis of covalently cross-linked receptors using several photosensitive CCK and gastrin probes of different peptide chain lengths demonstrated that in guinea-pig, CCK peptides bound to a 84-kDa component whereas in dog pancreas, CCK and gastrin peptides bound to three distinct molecular species (Mr approximately equal to 78,000, 45,000, 28,000). Performing cross-linking in the presence of 1 microM CCK indicated that a 45-kDa protein is the putative CCK/gastrin receptor in dog pancreas. Our results support the concept of heterogeneity of CCK/gastrin receptors.     

Evolution of new hormone function: loss and gain of a receptor

The vertebrate proglucagon gene encodes three glucagon-like sequences (glucagon, glucagon-like peptide-1 [GLP-1], and glucagon-like peptide 2 [GLP-2]) that have distinct functions in regulating metabolism in mammals. In contrast, glucagon and GLP-1 have similar physiological actions in fish, that of mammalian glucagon. We have identified sequences similar to receptors for proglucagon-derived peptides from the genomes of two fish (pufferfish and zebrafish), a frog (Xenopus tropicalis), and a bird (chicken). Phylogenetic analysis of the receptor sequences suggested an explanation for the divergent function of GLP-1 in fish and mammals. The phylogeny of our predicted and characterized receptors for proglucagon-derived peptides demonstrate that receptors for glucagon, GLP-1, and GLP-2 have an origin before the divergence of fish and mammals; however, fish have lost the gene encoding the GLP-1 class of receptors, and likely the incretin action of GLP-1. Receptors that bind GLP-1, but yield glucagon-like action, have been characterized in goldfish and zebrafish, and these sequences are most closely related to glucagon receptors. Both pufferfish and zebrafish have a second glucagon receptor-like gene that is most closely related to the characterized goldfish glucagon receptor. The phylogeny of glucagon receptor-like genes in fish indicates that a duplication of the glucagon receptor gene occurred on the ancestral fish lineage, and could explain the shared action of glucagon and GLP-1. We suggest that the binding specificity of one of the duplicated glucagon receptors has diverged, yielding receptors for GLP-1 and glucagon, but that ancestral downstream signaling has been maintained, resulting in both receptors retaining glucagon-stimulated downstream effects.     

Glucagon and the Glucagon Receptor: Merrifield Years at the Interface of Chemistry and Biology

The glucagon signaling system is a good model to investigate the chemical and structural requirements that dictate the interaction between a peptide hormone and its membrane-bound receptor and the cascade of events that lead to a physiological response. Secreted by pancreatic A cells, the primary target organ of glucagon is the liver where, together with insulin, it plays a central role in the maintenance of normal circulating glucose levels critical to the survival of the organism. The impetus for studying how glucagon interacts with its receptor is to gain insight into the mechanism of glucagon action in normal physiology as well as in diabetes mellitus. The principal approach towards this goal is to design and synthesize analogues of glucagon that will bind with high affinity to the glucagon receptor but will not activate it. These peptide analogues are expected to be potent antagonists of the hormone and will provide insight into the role of glucagon in diabetes. A second complementary approach is to investigate structure-function relationships in the glucagon receptor by site-directed mutagenesis and the biochemical and pharmacological characterization of mutant receptors. These studies will provide information about the peptide-binding site in the receptor and the residues that dictate ligand selectivity. A stable mammalian cell line that expresses human glucagon receptor at high-levels has been developed and should provide receptor protein for structural studies. An interdisciplinary approach combining chemical synthesis, molecular biology and biophysical methods is crucial for the conception of three-dimensional receptor models to be used in the rational design of glucagon antagonists for the management of diabetes. Dedicated to Bruce Merrifield.     

Biological activities of catfish glucagon and glucagon-like peptide

The ability of catfish glucagon and glucagon-like peptide to bind and activate mammalian glucagon receptors was investigated. Neither catfish peptide binds to glucagon receptors of rat liver, hypothalamus or pituitary. Neither stimulates adenylate cyclase activity in liver membranes. Catfish glucagon fails to activate adenylate cyclase in hypothalamic or pituitary membranes in contrast to mammalian glucagon. However, catfish glucagon-like peptide does stimulate hypothalamic and pituitary adenylate cyclase (EC50 approximately 1 pM) possibly through mammalian glucagon-like peptide receptors.     

Effects of neuromedin B and GRP-10 on gastrin and insulin release from cultured tumor cells of a malignant gastrinoma

A study relating to gastrin release from gastrinoma cells by neuromedin B and C-terminal decapeptide of gastrin releasing peptide (GRP-10) has not yet been reported. Therefore, we studied the effects of neuromedin B and GRP-10 on gastrin release from cultured dispersed cells prepared from both the primary tumor in the pancreas and the metastatic tumor in the liver from a case of malignant Zollinger-Ellison syndrome. Both the primary and metastatic tumors obtained by a curative operation contained similar concentrations of gastrin and glucagon, whereas the primary tumor contained 10 times more insulin than the metastatic tumor. Gastrin release from cultured cells of both tumors was suppressed by 0.1 and 10 nM neuromedin B and tended to be suppressed by 0.1-10 nM GRP-10. However, insulin release from cultured cells of the pancreatic tumor was stimulated by GRP-10, but not by neuromedin B. These results might suggest that receptor function for the bombesin family peptides is abnormal in gastrinoma cells in both primary and metastatic tumors, and that a major source of insulin secretary cells is the contaminated normal islet cells in the primary tumor.     

The effect of bariatric surgery on gastrointestinal and pancreatic peptide hormones

Bariatric surgery for obesity has proved to be an extremely effective method of promoting long-term weight reduction with additional beneficial metabolic effects, such as improved glucose tolerance and remission of type 2 diabetes. A range of bariatric procedures are in common use, including gastric banding, sleeve gastrectomy and the Roux-en-Y gastric bypass. Although the mechanisms underlying the efficacy of bariatric surgery are unclear, gastrointestinal and pancreatic peptides are thought to play an important role. The aim of this review is to summarise the effects of different bariatric surgery procedures upon gastrointestinal and pancreatic peptides, including ghrelin, gastrin, cholecystokinin (CCK), glucose-dependent insulinotropic hormone (GIP), glucagon-like peptide 1 (GLP-1), peptide YY (PYY), oxyntomodulin, insulin, glucagon and somatostatin.     

甲酰肽受体研究进展

趋化剂N-甲酰肽,如fMLF(N-甲酰甲硫氨酰-亮氨酰-苯丙氨酸)与受体结合后,能在炎症和免疫应急反应时募集嗜中性粒细胞游走和聚集在病灶处,对抗并清除病原微生物。近年来发现的许多结构各异的非N-甲酰肽配体(包括炎症早期出现的内源性多肽)均具有趋化和激活噬菌性白细胞的作用。这些研究进展拓展了我们对甲酰肽受体功能的认识,同时也提出一系列新问题,值得深入探讨。     

Regions of complementarity between the messenger RNAs for epidermal growth factor, transferrin, interleukin-2 and their respective receptors

Messenger (m)RNA sequences complementary to the mRNA sequences for the receptors to epidermal growth factor (EGF), interleukin-2 (IL-2) and transferrin (TF) were written out and compared for homologies with their ligands (EGF, IL-2 and TF, respectively). Highly significant amino acid and nucleotide homologies between the ligands and their appropriate receptor complements were detected in each case. For example, EGF and its receptor complement contained two homologous segments, each being six amino acids in length. When these segments were screened for matches against a protein sequence bank (3060 proteins and 616,748 test segments), only EGF contained either sequence. Similar results were obtained with IL-2 and TF. In each case, the homologous segments corresponded to complementary regions in the ligand binding portion of the receptor.     

Dose-response comparisons of canine plasma gastroenteropancreatic hormone responses to bombesin and the porcine gastrin-releasing peptide (GRP)

This study compares the potencies of the porcine gastrin-releasing peptide (pGRP) and bombesin, in causing elevations of canine plasma gastroenteropancreatic (GEP) levels. In the dose range 0-600 pmol . kg-1 . h-1, infusion of both peptides resulted in obvious dose-related elevations of plasma levels of gastrin, pancreatic polypeptide, enteroglucagon, immunoreactive pancreatic glucagon, and insulin. In this dose range, no significant difference in potency between the two peptides in elevating plasma levels of the above hormones was observed. The results of this study, demonstrating equimolar potency of pGRP and bombesin, are in contrast to previous studies reporting that pGRP was less potent than bombesin in causing certain bioactivities in the rat following intracranial administration of the two peptides.     

Paradoxical antagonism of PACAP receptor signaling by VIP in Xenopus oocytes via the type-C natriuretic peptide receptor

Atrial natriuretic peptide (ANP) and the closely-related peptides BNP and CNP are highly conserved cardiovascular hormones. They bind to single transmembrane-spanning receptors, triggering receptor-intrinsic guanylyl cyclase activity. The "truncated" type-C natriuretic peptide receptor (NPR-C) has long been called a clearance receptor because it lacks the intracellular guanylyl cyclase domain, though data suggest it might negatively couple to adenylyl cyclase via G(i). Here we report the molecular cloning and characterization of the Xenopus laevis type-C natriuretic peptide receptor (XNPR-C). Analysis confirms the presence of a short intracellular C-terminus, as well as a high similarity to fish and mammalian NPR-C. Injection of XNPR-C mRNA into Xenopus oocytes resulted in expression of high affinity [(125)I]ANP binding sites that were competitively and completely displaced by natriuretic analogs and the unrelated neuropeptide vasoactive intestinal peptide (VIP). Measurement of cAMP levels in mRNA-injected oocytes revealed that XNPR-C is negatively coupled to adenylyl cyclase in a pertussis toxin-sensitive manner. When XNPR-C was co-expressed with PAC(1) receptors for pituitary adenylyl cyclase-activating polypeptide (PACAP), VIP and natriuretic peptides counteracted the cAMP induction by PACAP. These results suggest that VIP and natriuretic peptides can potentially modulate the action of PACAP in cells where these receptors are co-expressed.     

Rabies virus binding to an acetylcholine receptor alpha-subunit peptide

The binding of 125I-labeled rabies virus to a synthetic peptide comprising residues 173-204 of the alpha 1-subunit of the nicotinic acetylcholine receptor was investigated. Binding of rabies virus to the receptor peptide was dependent on pH, could be competed with by unlabeled homologous virus particles, and was saturable. Synthetic peptides of snake venom, curaremimetic neurotoxins and of the structurally similar segment of the rabies virus glycoprotein, were effective in competing with labeled virus binding to the receptor peptide at micromolar concentrations. Similarly, synthetic peptides of the binding domain on the acetylcholine receptor competed for binding. These findings suggest that both rabies virus and neurotoxins bind to residues 173-204 of the alpha 1-subunit of the acetylcholine receptor. Competition studies with shorter alpha-subunit peptides within this region indicate that the highest affinity virus binding determinants are located within residues 179-192. A rat nerve alpha 3-subunit peptide, that does not bind alpha-bungarotoxin, inhibited binding of virus to the alpha 1 peptide, suggesting that rabies binds to neuronal nicotinic acetylcholine receptors. These studies indicate that synthetic peptides of the glycoprotein binding domain and of the receptor binding domain may represent useful antiviral agents by targeting the recognition event between the viral attachment protein and the host cell receptor, and inhibiting attachment of virus to the receptor.     

Insulin-binding peptide. Design and characterization

The design and characterization of a six-amino acid-containing peptide that binds insulin is described. The amino acid sequence of the insulin-binding peptide (IBP) was determined from the strand of DNA complementary to the strand of DNA coding for the insulin molecule in the domain of the insulin monomer believed to interact with the insulin receptor. The IBP (Cys-Val-Glu-Glu-Ala-Ser) binds specifically to insulin in a saturable manner with a Kd of 3 nM. This binding process is time dependent and slightly temperature dependent, and the peptide appears to interact with insulin near the carboxyl terminus of the B-chain of insulin. Incubation of insulin with the peptide decreases insulin binding to the insulin receptor by 50%, with no effect on the affinity of insulin for the receptor and no effect on cellular insulin-stimulated deoxyglucose uptake. A polyclonal antibody produced against the IBP will inhibit specific insulin binding to intact cells by approximately 50%, with no effects on insulin-stimulated glucose uptake. From this data, we suggest that there are at least two domains of the insulin molecule through which it interacts with its receptor, the "binding region" of insulin, which is the domain blocked by the IBP, and the "message region" of insulin, through which insulin not only binds to the receptor, but also generates the cellular signal.