scholarly journals Comparison of Inhibitor and Substrate Selectivity between Rodent and Human Vascular Adhesion Protein-1

2020 ◽  
Vol 2020 ◽  
pp. 1-9 ◽  
Author(s):  
Ryo Kubota ◽  
Michael J. Reid ◽  
Kuo Lee Lieu ◽  
Mark Orme ◽  
Christine Diamond ◽  
...  
Keyword(s):  
Amine Oxidase ◽  
Leukocyte Adhesion ◽  
Primary Amines ◽  
Oxidative Deamination ◽  
Adhesion Protein ◽  
Common Marker ◽  
Small Primary ◽  
Vascular Adhesion ◽  
Vascular Adhesion Protein 1

Vascular adhesion protein-1 (VAP-1) is an ectoenzyme that functions as a copper-containing amine oxidase and is involved in leukocyte adhesion at sites of inflammation. Inhibition of VAP-1 oxidative deamination has become an attractive target for anti-inflammatory therapy with demonstrated efficacy in rodent models of inflammation. A previous comparison of purified recombinant VAP-1 from mouse, rat, monkey, and human gene sequences predicted that rodent VAP-1 would have higher affinity for smaller hydrophilic substrates/inhibitors because of its narrower and more hydrophilic active site channel. An optimized in vitro oxidative deamination fluorescence assay with benzylamine (BA) was used to compare inhibition of five known inhibitors in recombinant mouse, rat, and human VAP-1. Human VAP-1 was more sensitive compared to rat or mouse VAP-1 (lowest IC50 concentration) to semicarbazide but was least sensitive to hydralazine and LJP-1207. Hydralazine had a lower IC50 in rats compared to humans, although not significant. However, the IC50 of hydralazine was significantly higher in the rat compared to mouse VAP-1. The larger hydrophobic compounds from Astellas (compound 35c) and Boehringer Ingelheim (PXS-4728A) were hypothesized to have higher binding affinity for human VAP-1 compared to rodent VAP-1 since the channel in human VAP-1 is larger and more hydrophobic than that in rodent VAP-1. Although the sensitivity of these two inhibitors was the lowest in the mouse enzyme, we found no significant differences between mouse, rat, and human VAP-1. Michaelis-Menten kinetics of the small primary amines phenylethylamine and tyramine were also compared to the common marker substrate BA demonstrating that BA had the highest affinity among the substrates. Rat VAP-1 had the highest affinity for all three substrates and mouse VAP-1 had intermediate affinity for BA and phenylethylamine, but tyramine was not a substrate for mouse VAP-1 under these assay conditions. These results suggest that comparing oxidative deamination in mouse and rat VAP-1 may be important if using these species for preclinical efficacy models.

scholarly journals Siglec-9 is a novel leukocyte ligand for vascular adhesion protein-1 and can be used in PET imaging of inflammation and cancer

Blood ◽  
2011 ◽  
Vol 118 (13) ◽  
pp. 3725-3733 ◽  
Author(s):  
Kristiina Aalto ◽  
Anu Autio ◽  
Elina A. Kiss ◽  
Kati Elima ◽  
Yvonne Nymalm ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Primary Amines ◽  
Dependent Manner ◽  
Binding Assays ◽  
Adhesion Protein ◽  
Interaction Sites ◽  
Positron Emission ◽  
Vascular Adhesion ◽  
Vascular Adhesion Protein 1

Abstract Leukocyte migration to sites of inflammation is regulated by several endothelial adhesion molecules. Vascular adhesion protein-1 (VAP-1) is unique among the homing-associated molecules as it is both an enzyme that oxidizes primary amines and an adhesin. Although granulocytes can bind to endothelium via a VAP-1–dependent manner, the counter-receptor(s) on this leukocyte population is(are) not known. Here we used a phage display approach and identified Siglec-9 as a candidate ligand on granulocytes. The binding between Siglec-9 and VAP-1 was confirmed by in vitro and ex vivo adhesion assays. The interaction sites between VAP-1 and Siglec-9 were identified by molecular modeling and confirmed by further binding assays with mutated proteins. Although the binding takes place in the enzymatic groove of VAP-1, it is only partially dependent on the enzymatic activity of VAP-1. In positron emission tomography, the 68Gallium-labeled peptide of Siglec-9 specifically detected VAP-1 in vasculature at sites of inflammation and cancer. Thus, the peptide binding to the enzymatic groove of VAP-1 can be used for imaging conditions, such as inflammation and cancer.

Granulocyte transmigration through the endothelium is regulated by the oxidase activity of vascular adhesion protein-1 (VAP-1)

Blood ◽  
2004 ◽  
Vol 103 (9) ◽  
pp. 3388-3395 ◽  
Author(s):  
Kaisa Koskinen ◽  
Petri J. Vainio ◽  
David J. Smith ◽  
Marjo Pihlavisto ◽  
Seppo Ylä-Herttuala ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Oxidase Activity ◽  
Enzyme Inhibitors ◽  
Amine Oxidase ◽  
Laminar Shear Stress ◽  
Adhesion Protein ◽  
Vascular Adhesion ◽  
Vascular Adhesion Protein 1

Abstract Polymorphonuclear leukocytes (PMNs) migrate from the blood into areas of inflammation by binding to the endothelial cells of blood vessels via adhesion molecules. Vascular adhesion protein-1 (VAP-1) is one of the molecules mediating leukocyte-endothelial cell interactions. It is also an endothelial cell-surface enzyme (amine oxidase) that produces reactive oxygen species during the catalytic reaction. To study the role of the enzymatic activity of VAP-1 in PMN extravasation, we used an enzymatically inactive VAP-1 mutant, specific amine oxidase inhibitors (including a novel small molecule compound), and anti-VAP-1 antibodies in several flow-dependent models. The enzyme inhibitors diminished PMN rolling on and transmigration through human endothelial cells under conditions of laminar shear stress in vitro. Notably, the enzyme inactivating point mutation abolished the capacity of VAP-1 to mediate transmigration. Moreover, the new VAP-1 inhibitor effectively prevented the extravasation of PMNs in an animal model of inflammation. These data show that the oxidase activity of VAP-1 controls PMN exit from the blood during the relatively poorly understood transmigration step. (Blood. 2004;103:3388-3395)

scholarly journals Human Siglec-10 can bind to vascular adhesion protein-1 and serves as its substrate

Blood ◽  
2009 ◽  
Vol 114 (26) ◽  
pp. 5385-5392 ◽  
Author(s):  
Elina Kivi ◽  
Kati Elima ◽  
Kristiina Aalto ◽  
Yvonne Nymalm ◽  
Kaisa Auvinen ◽  
...  
Keyword(s):  
Primary Amines ◽  
Leukocyte Trafficking ◽  
Hydrogen Peroxide Production ◽  
Lymphocyte Adhesion ◽  
Adhesion Protein ◽  
Inflammatory Microenvironment ◽  
Inflamed Endothelium ◽  
End Products ◽  
Vascular Adhesion ◽  
Vascular Adhesion Protein 1

Abstract Leukocytes migrate from the blood into areas of inflammation by interacting with various adhesion molecules on endothelial cells. Vascular adhesion protein-1 (VAP-1) is a glycoprotein expressed on inflamed endothelium where it plays a dual role: it is both an enzyme that oxidizes primary amines and an adhesin that is involved in leukocyte trafficking to sites of inflammation. Although VAP-1 was identified more than 15 years ago, the counterreceptor(s) for VAP-1 on leukocytes has remained unknown. Here we have identified Siglec-10 as a leukocyte ligand for VAP-1 using phage display screenings. The binding between Siglec-10 and VAP-1 was verified by different adhesion assays, and this interaction was also consistent with molecular modeling. Moreover, the interaction between Siglec-10 and VAP-1 led to increased hydrogen peroxide production, indicating that Siglec-10 serves as a substrate for VAP-1. Thus, the Siglec-10–VAP-1 interaction seems to mediate lymphocyte adhesion to endothelium and has the potential to modify the inflammatory microenvironment via the enzymatic end products.

scholarly journals Zinc-α2-Glycoprotein Is An Inhibitor Of Amine Oxidase Copper-Containing 3

2019 ◽  
Author(s):  
Matthias Romauch
Keyword(s):  
Amine Oxidase ◽  
Current Knowledge ◽  
Primary Amines ◽  
Adhesion Protein ◽  
Diagnosis And Prognosis ◽  
Fat Mobilization ◽  
Clinical Biomarker ◽  
Progressive Weight Loss ◽  
Inhibitory Potential ◽  
Vascular Adhesion

AbstractZinc-alpha2-glycoprotein (ZAG) is a major plasma protein whose levels increase in chronic energy-demanding diseases and thus serves as an important clinical biomarker in the diagnosis and prognosis of the development of cachexia. Current knowledge suggests that ZAG mediates progressive weight loss through β-adrenergic signaling in adipocytes, resulting in the activation of lipolysis and fat mobilization. Here, through crosslinking experiments, amine oxidase copper-containing 3 (AOC3) is identified as a novel ZAG binding partner. AOC3 – also known as vascular adhesion protein 1 (VAP-1) and semicarbazide sensitive amine oxidase (SSAO) – deaminates primary amines, thereby generating the corresponding aldehyde, H2O2 and HN3. It is an ectoenzyme largely expressed by adipocytes and induced in endothelial cells during inflammation. Extravasation of immune cells depends on amine oxidase activity and AOC3-derived H2O2 has an insulinogenic effect. The observations described here suggest that ZAG acts as an allosteric inhibitor of AOC3 and interferes with the associated pro-inflammatory and anti-lipolytic functions. Thus, inhibition of the deamination of lipolytic hormone octopamine by AOC3 represents a novel mechanism by which ZAG might stimulate lipolysis. Furthermore, experiments involving overexpression of recombinant ZAG reveal that its glycosylation is co-regulated by oxygen availability and that the pattern of glycosylation affects its inhibitory potential. The newly identified protein interaction between AOC3 and ZAG highlights a previously unknown functional relationship, which may be relevant to inflammation, energy metabolism and the development of cachexia.

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