Novel carbon skeletons activate human NicotinAMide Phosphoribosyl Transferase (NAMPT) enzyme in biochemical assay.

Nicotinamide adenine dinucleotide (NAD) is a central molecule in cellular metabolism that has been implicated in human health, the aging process, and an array of human diseases. NAD is well known as an electron storage molecule, cycling between NAD and the reduced NADH. In addition, NAD is cleaved into nicotinamide and Adenine diphosphate ribose by NAD-consuming enzymes such as sirtuins, PARPs and CD38.

O-fucosylation of thrombospondin type I repeats is dispensable for trafficking thrombospondin 1 to platelet secretory granules.

Thrombospondin 1 (THBS1) is a secreted extracellular matrix glycoprotein that regulates a variety of cellular and physiological processes. THBS1's diverse functions are attributed to interactions between the modular domains of THBS1 with an array of proteins found in the extracellular matrix. THBS1's three Thrombospondin type 1 repeats (TSRs) are modified with O-linked glucose-fucose disaccharide and C-mannose.

O-fucosylation stabilizes the TSR3 motif in thrombospondin-1 by interacting with nearby amino acids and protecting a disulfide bond.

Thrombospondin type-1 repeats (TSRs) are small protein motifs containing six conserved cysteines forming three disulfide bonds that can be modified with an O-linked fucose. Protein O-fucosyltransferase 2 (POFUT2) catalyzes the addition of O-fucose to TSRs containing the appropriate consensus sequence, and the O-fucose modification can be elongated to a Glucose-Fucose disaccharide with the addition of glucose by β3-glucosyltransferase (B3GLCT). Elimination of Pofut2 in mice results in embryonic lethality in mice, highlighting the biological significance of O-fucose modification on TSRs.

O-fucosylation of thrombospondin type 1 repeats is essential for ECM remodeling and signaling during bone development.

Many extracellular matrix (ECM) associated proteins that influence ECM properties have Thrombospondin type 1 repeats (TSRs) which are modified with O-linked fucose. The O-fucose is added in the endoplasmic reticulum to folded TSRs by the enzyme Protein O-fucosyltransferase-2 (POFUT2) and is proposed to promote efficient trafficking of substrates. The importance of this modification for function of TSR-proteins is underscored by the early embryonic lethality of mouse embryos lacking Pofut2.

Hydrocephalus in mouse B3glct mutants is likely caused by defects in multiple B3GLCT substrates in ependymal cells and subcommissural organ.

Peters plus syndrome, characterized by defects in eye and skeletal development with isolated cases of ventriculomegaly/hydrocephalus, is caused by mutations in the β3-glucosyltransferase (B3GLCT) gene. In the endoplasmic reticulum, B3GLCT adds glucose to O-linked fucose on properly folded thrombospondin type 1 repeats (TSRs). The resulting glucose-fucose disaccharide is proposed to stabilize the TSR fold and promote secretion of B3GLCT substrates, with some substrates more sensitive than others to loss of glucose.

Fucosylation of ADAMTSL2 is required for secretion and is impacted by geleophysic dysplasia-causing mutations.

mutations cause an autosomal recessive connective tissue disorder, geleophysic dysplasia 1 (GPHYSD1), which is characterized by short stature, small hands and feet, and cardiac defects. ADAMTSL2 is a matricellular protein previously shown to interact with latent transforming growth factor-β binding protein 1 and influence assembly of fibrillin 1 microfibrils. ADAMTSL2 contains seven thrombospondin type-1 repeats (TSRs), six of which contain the consensus sequence for -fucosylation by protein fucosyltransferase 2 (POFUT2).

ADAMTS9 and ADAMTS20 are differentially affected by loss of B3GLCT in mouse model of Peters plus syndrome.

Peters plus syndrome (MIM #261540 PTRPLS), characterized by defects in eye development, prominent forehead, hypertelorism, short stature and brachydactyly, is caused by mutations in the β3-glucosyltransferase (B3GLCT) gene. Protein O-fucosyltransferase 2 (POFUT2) and B3GLCT work sequentially to add an O-linked glucose β1-3fucose disaccharide to properly folded thrombospondin type 1 repeats (TSRs). Forty-nine proteins are predicted to be modified by POFUT2, and nearly half are members of the ADAMTS superfamily.

Analyzing the Effects of O-Fucosylation on Secretion of ADAMTS Proteins Using Cell-Based Assays.

Metalloproteinases of the ADAMTS (a disintegrin and metalloproteinase with thrombospondin type 1 repeats) superfamily are extensively modified with glycan moieties. Glycosylation occurs as these enzymes are trafficked through the endoplasmic reticulum (ER) and Golgi apparatus on their way to the extracellular space and includes N-linked glycosylation, O-linked fucosylation and C-linked mannosylation. This chapter focuses on O-linked fucose, which is added to properly folded thrombospondin type 1 repeats (TSRs) in the ER by protein O-fucosyltransferase 2 (POFUT2) and elongated to a Glucoseβ1-3Fucose disaccharide by β3-glucosyltransferase (B3GLCT).

Structure of the IFNγ receptor complex guides design of biased agonists.

The cytokine interferon-γ (IFNγ) is a central coordinator of innate and adaptive immunity, but its highly pleiotropic actions have diminished its prospects for use as an immunotherapeutic agent. Here, we took a structure-based approach to decoupling IFNγ pleiotropy. We engineered an affinity-enhanced variant of the ligand-binding chain of the IFNγ receptor IFNγR1, which enabled us to determine the crystal structure of the complete hexameric (2:2:2) IFNγ-IFNγR1-IFNγR2 signalling complex at 3.

Variant in human POFUT1 reduces enzymatic activity and likely causes a recessive microcephaly, global developmental delay with cardiac and vascular features.

Protein O-fucosyltransferase-1 (POFUT1) adds O-fucose monosaccharides to epidermal growth factor-like (EGF) repeats found on approximately 100 mammalian proteins, including Notch receptors. Haploinsufficiency of POFUT1 has been linked to adult-onset Dowling Degos Disease (DDD) with hyperpigmentation defects. Homozygous deletion of mouse Pofut1 results in embryonic lethality with severe Notch-like phenotypes including defects in somitogenesis, cardiogenesis, vasculogenesis and neurogenesis, but the extent to which POFUT1 is required for normal human development is not yet understood.

Unusual life cycle and impact on microfibril assembly of ADAMTS17, a secreted metalloprotease mutated in genetic eye disease.

Secreted metalloproteases have diverse roles in the formation, remodeling, and the destruction of extracellular matrix. Recessive mutations in the secreted metalloprotease ADAMTS17 cause ectopia lentis and short stature in humans with Weill-Marchesani-like syndrome and primary open angle glaucoma and ectopia lentis in dogs. Little is known about this protease or its connection to fibrillin microfibrils, whose major component, fibrillin-1, is genetically associated with ectopia lentis and alterations in height.