Turning the spotlight on the oligosaccharide chain of GM1 ganglioside

Abstract

It is well over a century that glycosphingolipids are matter of interest in different fields of research. The hydrophilic oligosaccharide and the lipid moiety, the ceramide, both or separately have been considered in different moments as the crucial portion of the molecule, responsible for the role played by the glycosphingolipids associated to the plasma-membranes or to any other subcellular fraction. Glycosphingolipids are a family of compounds characterized by thousands of structures differing in both the oligosaccharide and the ceramide moieties, but among them, the nervous system monosialylated glycosphingolipid GM1, belonging to the group of gangliosides, has gained particular attention by a multitude of Scientists. In recent years, a series of studies have been conducted on the functional roles played by the hydrophilic part of GM1, its oligosaccharide, that we have named "OligoGM1". These studies allowed to shed new light on the mechanisms underlying the properties of GM1 defining the role of the OligoGM1 in determining precise interactions with membrane proteins instrumental for the neuronal functions, leaving to the ceramide the role of correctly positioning the GM1 in the membrane crucial for the oligosaccharide-protein interactions. In this review we aim to report the recent studies on the cascade of events modulated by OligoGM1, as the bioactive portion of GM1, to support neuronal differentiation and trophism together with preclinical studies on its potential to modify the progression of Parkinson's disease.

Keywords: Drug development; GM1 oligosaccharide; Neurodegeneration; Neurodifferentiation; Neuroprotection; Parkinson’s disease.

Fig. 1

Structure of ganglioside GM1, II³Neu5AcGg₄-Cer

Fig. 2

OligoGM1 synthesis strategies. a Scheme of the chemical synthesis of OligoGM1. GM1 undergoes to ozonolytic (O₃) process in methanol (i) generating a desphingosine aldehyde product and a miristic aldehyde (ii), followed by alkaline hydrolysis in triethylamine (Et₃N) (iii) releasing OlgoGM1; b Scheme of the chemical synthesis of tritium-labeled and photoactivable OligoGM1. [³H]GM1 undergoes ozonolysis (i) in methanol followed by triethylamine alkaline hydrolysis (ii) releasing [³H]OligoGM1. The latter is subjected to the amination process (iii) and the [³H]OligoGM1-NH₂ is azide labeled (iv) with 2-nitro-fluorophenylazide in dimethylformamide (DMF) and tributiylamine (Bu₃N) in dimethylsulfoxide (DMSO) to obtain [³H]OligoGM1-N₃. c OligoGM1 biosynthesis in engineered Escherichia coli. Lactose and sialic acid (Neu5Ac) are internalized by the specific permeases LacY and NanT but cannot be degraded because of β-galactosidase (LacZ) and aldolase (NanA) deletion. CMP-Neu5Ac synthase activates Neu5Ac into CMP-Neu5Ac which is transferred onto lactose by α2,3-sialyltransferase (encoded by Lst), to form II³αNeu5Ac-Lac (sialyllactose, OligoGM3). The use of the endogenous pool of UDP-GalNAc produced by the recombinant UDP-GlcNAc C4 epimerase (WbpP) allows β1,4-GalNAc transferase (CgtA) to catalyze the glycosylation of sialyllactose to form II³αNeu5Ac-Gg₃ (OligoGM2). This compound is substrate for the β1,3-galactosyltransferase (CgtB) to yield II³αNeu5Ac-Gg₄ (OligoGM1). CTP, cytidine triphosphate; Ppi, inorganic pyrophosphate. Oligosaccharide sugar code is according to Varki et al. [36]

Fig. 2

OligoGM1 synthesis strategies. a Scheme of the chemical synthesis of OligoGM1. GM1 undergoes to ozonolytic (O₃) process in methanol (i) generating a desphingosine aldehyde product and a miristic aldehyde (ii), followed by alkaline hydrolysis in triethylamine (Et₃N) (iii) releasing OlgoGM1; b Scheme of the chemical synthesis of tritium-labeled and photoactivable OligoGM1. [³H]GM1 undergoes ozonolysis (i) in methanol followed by triethylamine alkaline hydrolysis (ii) releasing [³H]OligoGM1. The latter is subjected to the amination process (iii) and the [³H]OligoGM1-NH₂ is azide labeled (iv) with 2-nitro-fluorophenylazide in dimethylformamide (DMF) and tributiylamine (Bu₃N) in dimethylsulfoxide (DMSO) to obtain [³H]OligoGM1-N₃. c OligoGM1 biosynthesis in engineered Escherichia coli. Lactose and sialic acid (Neu5Ac) are internalized by the specific permeases LacY and NanT but cannot be degraded because of β-galactosidase (LacZ) and aldolase (NanA) deletion. CMP-Neu5Ac synthase activates Neu5Ac into CMP-Neu5Ac which is transferred onto lactose by α2,3-sialyltransferase (encoded by Lst), to form II³αNeu5Ac-Lac (sialyllactose, OligoGM3). The use of the endogenous pool of UDP-GalNAc produced by the recombinant UDP-GlcNAc C4 epimerase (WbpP) allows β1,4-GalNAc transferase (CgtA) to catalyze the glycosylation of sialyllactose to form II³αNeu5Ac-Gg₃ (OligoGM2). This compound is substrate for the β1,3-galactosyltransferase (CgtB) to yield II³αNeu5Ac-Gg₄ (OligoGM1). CTP, cytidine triphosphate; Ppi, inorganic pyrophosphate. Oligosaccharide sugar code is according to Varki et al. [36]

Fig. 2

OligoGM1 synthesis strategies. a Scheme of the chemical synthesis of OligoGM1. GM1 undergoes to ozonolytic (O₃) process in methanol (i) generating a desphingosine aldehyde product and a miristic aldehyde (ii), followed by alkaline hydrolysis in triethylamine (Et₃N) (iii) releasing OlgoGM1; b Scheme of the chemical synthesis of tritium-labeled and photoactivable OligoGM1. [³H]GM1 undergoes ozonolysis (i) in methanol followed by triethylamine alkaline hydrolysis (ii) releasing [³H]OligoGM1. The latter is subjected to the amination process (iii) and the [³H]OligoGM1-NH₂ is azide labeled (iv) with 2-nitro-fluorophenylazide in dimethylformamide (DMF) and tributiylamine (Bu₃N) in dimethylsulfoxide (DMSO) to obtain [³H]OligoGM1-N₃. c OligoGM1 biosynthesis in engineered Escherichia coli. Lactose and sialic acid (Neu5Ac) are internalized by the specific permeases LacY and NanT but cannot be degraded because of β-galactosidase (LacZ) and aldolase (NanA) deletion. CMP-Neu5Ac synthase activates Neu5Ac into CMP-Neu5Ac which is transferred onto lactose by α2,3-sialyltransferase (encoded by Lst), to form II³αNeu5Ac-Lac (sialyllactose, OligoGM3). The use of the endogenous pool of UDP-GalNAc produced by the recombinant UDP-GlcNAc C4 epimerase (WbpP) allows β1,4-GalNAc transferase (CgtA) to catalyze the glycosylation of sialyllactose to form II³αNeu5Ac-Gg₃ (OligoGM2). This compound is substrate for the β1,3-galactosyltransferase (CgtB) to yield II³αNeu5Ac-Gg₄ (OligoGM1). CTP, cytidine triphosphate; Ppi, inorganic pyrophosphate. Oligosaccharide sugar code is according to Varki et al. [36]

Fig. 3

Neuritogenic potential of GM1, OligoGM1 and isolated carbohydrate’s components in N2a cells. a The symbolic structure of tested compounds according to [36], whose specific names are reported in the table below. b The effectiveness of the compound in inducing neuritic growth is represented by the + symbol. The symbol - indicates that the compound fails to induce neuritogenesis

Fig. 4

Diagram of the proposed mechanism for OligoGM1 modulatory activities in N2a cells. At cell surface, OligoGM1 is able to directly bind and stabilize the TrkA-NGF complex triggering the hyperphosphorylation of the receptor at Tyr490 and the downstream MAPK pathway. The activated cascade stimulates several cellular events including the calcium signalling, the differentiation in neuron-like cells, the protection against neurotoxic stimuli (i.e. MPTP), the enhancement of mitochondrial bioenergetics. This image is updated from [28, 29, 31, 33]. GM1 saccharide representation is according to [36]. ERK, extracellular signal regulated protein kinases 1 and 2; Grb2, growth factor receptor bound protein 2; Gab1, Grb2-associated binder-1; RAS, GTP-binding protein; RAF, serine/threonine kinase; SHC, transforming protein 1; SOS, son of sevenless; p38 Mitogen-activated protein kinase, PLCγ, Phospholipase Cγ; PIP2, Phosphatidylinositol 4,5-bisphosphate; DAG, Diacylglycerol; IP₃, Inositol trisphosphate; PKC, Protein kinase C

Fig. 5

Molecular docking analyses of TrkA-NGF-OligoGM1 complex. a Diagram depicting the TrkA-NGF complex at cell surface and representation showing the TrkA-NGF interaction in absence (left) or in presence (right) of OligoGM1 as crystallographic complex. The binding energy associated to the TrkA-NGF complex is reduced from − 7 kcal/mol to -12 kcal/mol when OligoGM1 is present into the complex TrkA receptor is represented in magenta ribbons; NGF molecules in orange ribbons; OligoGM1 using space-filling model. b Representation of bonds between the extracellular residues of TrkA, NGF and OligoGM1

Fig. 6

Structure of LIGA20

Fig. 7

Schematic representation of OligoGM1-mediated neuroprotection in PD. On the left panel is shown the in vitro neuroprotective activity of OligoGM1 against MPTP neurotoxin, a widely accepted PD model. a MPTP exposure determines a deep mitochondrial impairment with a strong production of ROS that finally leads to cell death. c The administration of OligoGM1 strongly reduces the mitochondrial ROS content counteracting the MPTP toxicity and enhancing the cell survival [29]. On the right panel is represented the OligoGM1 capability to recover the PD features of GM1 deficient (B4galnt1^+/−) mice. b The reduced GM1 content triggers a profound motor impairment due to the loss of substantia nigra pars compacta (SNpc) dopaminergic neurons lesioned by α-syn oligomers accumulation and to the consequent dopamine content reduction. d The chronic injection of OligoGM1 is able to completely counteract the PD phenotype: recovery of motor symptoms, rescue of dopaminergic neurons, dopamine levels and clearance of α-syn aggregates [94]. Sugar representation is according to [36]