Cloning and functional characterization of a membrane-bound proinflamatory protein in Helicobacter pylori (HP-MBP1)

ABSTRACT Vibrio vulnificus is a Gram-negative bacterium that causes a fatal septicemia. One of its virulence factors is a membrane-bound lipoprotein, IlpA, which can induce cytokine production in human immune cells. In the present study, the role of IlpA as an adhesion molecule was investigated. An ilpA-deleted V. vulnificus mutant showed significantly decreased adherence to INT-407 human intestinal epithelial cells, which in turn resulted in reduced cytotoxicity. The ΔilpA mutant recovered the adherence ability of the wild type by complementation in trans with the intact ilpA gene. In addition, pretreatment of V. vulnificus with anti-IlpA polyclonal antibodies resulted in a significant reduction of bacterial adherence. To localize the domain of IlpA required for cytoadherence, three truncated recombinant IlpA polypeptides were constructed and tested for the ability to adhere to human cells by a ligand-binding immunoblot assay and fluorescence microscopy. The polypeptide containing the carboxy (C)-terminal hydrophilic domain exhibited direct binding to INT-407 cells. Therefore, the C-terminal domain of IlpA allows this protein to be an adhesion molecule of V. vulnificus.

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Functional Characterization of Colon-Cancer-Associated Variants in ADAM17 Affecting the Catalytic Domain

Biomedicines ◽

10.3390/biomedicines8110463 ◽

2020 ◽

Vol 8 (11) ◽

pp. 463

Author(s):

Jan Philipp Dobert ◽

Anne-Sophie Cabron ◽

Philipp Arnold ◽

Egor Pavlenko ◽

Stefan Rose-John ◽

...

Keyword(s):

Catalytic Domain ◽

Functional Characterization ◽

Tumor Development ◽

Point Mutations ◽

Modern World ◽

Protective Effects ◽

Tissue Samples ◽

Membrane Bound

Although extensively investigated, cancer is still one of the most devastating and lethal diseases in the modern world. Among different types, colorectal cancer (CRC) is most prevalent and mortal, making it an important subject of research. The metalloprotease ADAM17 has been implicated in the development of CRC due to its involvement in signaling pathways related to inflammation and cell proliferation. ADAM17 is capable of releasing membrane-bound proteins from the cell surface in a process called shedding. A deficiency of ADAM17 activity has been previously shown to have protective effects against CRC in mice, while an upregulation of ADAM17 activity is suspected to facilitate tumor development. In this study, we characterize ADAM17 variants found in tissue samples of cancer patients in overexpression studies. We here focus on point mutations identified within the catalytic domain of ADAM17 and could show a functional dysregulation of the CRC-associated variants. Since the catalytic domain of ADAM17 is the only region structurally determined by crystallography, we study the effect of each point mutation not only to learn more about the role of ADAM17 in cancer, but also to investigate the structure–function relationships of the metalloprotease.

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Mechanism and Inhibition of Matrix Metalloproteinases

Current Medicinal Chemistry ◽

10.2174/0929867325666180326163523 ◽

2019 ◽

Vol 26 (15) ◽

pp. 2609-2633 ◽

Cited By ~ 8

Author(s):

Linda Cerofolini ◽

Marco Fragai ◽

Claudio Luchinat

Keyword(s):

Matrix Metalloproteinases ◽

Functional Characterization ◽

Structural Data ◽

Therapeutic Strategies ◽

Outer Cell ◽

Drug Candidates ◽

The Family ◽

Outer Cell Membrane ◽

Membrane Bound

Matrix metalloproteinases hydrolyze proteins and glycoproteins forming the extracellular matrix, cytokines and growth factors released in the extracellular space, and membrane-bound receptors on the outer cell membrane. The pathological relevance of MMPs has prompted the structural and functional characterization of these enzymes and the development of synthetic inhibitors as possible drug candidates. Recent studies have provided a better understanding of the substrate preference of the different members of the family, and structural data on the mechanism by which these enzymes hydrolyze the substrates. Here, we report the recent advancements in the understanding of the mechanism of collagenolysis and elastolysis, and we discuss the perspectives of new therapeutic strategies for targeting MMPs.

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