Characterization of Microfibrillar-associated Protein 4 (MFAP4) as a Tropoelastin- and Fibrillin-binding Protein Involved in Elastic Fiber Formation

Bartosz Pilecki; Anne T Holm; Anders Schlosser; Jesper B Moeller; Alexander P Wohl; Alexandra V Zuk; Stefanie E Heumüller; Russell Wallis; Soren K Moestrup; Gerhard Sengle; Uffe Holmskov; Grith L Sorensen

doi:10.1074/jbc.M115.681775

Characterization of Microfibrillar-associated Protein 4 (MFAP4) as a Tropoelastin- and Fibrillin-binding Protein Involved in Elastic Fiber Formation

J Biol Chem. 2016 Jan 15;291(3):1103-14. doi: 10.1074/jbc.M115.681775. Epub 2015 Nov 24.

Affiliations

¹ From the Department of Cancer and Inflammation Research, Institute of Molecular Medicine, Faculty of Health Sciences, University of Southern Denmark, 5000 Odense C, Denmark.
² the Center for Biochemistry, Faculty of Medicine and.
³ the Center for Biochemistry, Faculty of Medicine and the Center for Molecular Medicine Cologne, University of Cologne, 50931 Cologne, Germany.
⁴ the Department of Infection, Immunity and Inflammation, and Department of Molecular and Cell Biology, University of Leicester, Leicester LE1 9HN, United Kingdom, and.
⁵ From the Department of Cancer and Inflammation Research, Institute of Molecular Medicine, Faculty of Health Sciences, University of Southern Denmark, 5000 Odense C, Denmark, the Department of Clinical Biochemistry and Pharmacology, Odense University Hospital, 5000 Odense C, Denmark.
⁶ From the Department of Cancer and Inflammation Research, Institute of Molecular Medicine, Faculty of Health Sciences, University of Southern Denmark, 5000 Odense C, Denmark, glsorensen@health.sdu.dk.

Abstract

MFAP4 (microfibrillar-associated protein 4) is an extracellular glycoprotein found in elastic fibers without a clearly defined role in elastic fiber assembly. In the present study, we characterized molecular interactions between MFAP4 and elastic fiber components. We established that MFAP4 primarily assembles into trimeric and hexameric structures of homodimers. Binding analysis revealed that MFAP4 specifically binds tropoelastin and fibrillin-1 and -2, as well as the elastin cross-linking amino acid desmosine, and that it co-localizes with fibrillin-1-positive fibers in vivo. Site-directed mutagenesis disclosed residues Phe(241) and Ser(203) in MFAP4 as being crucial for type I collagen, elastin, and tropoelastin binding. Furthermore, we found that MFAP4 actively promotes tropoelastin self-assembly. In conclusion, our data identify MFAP4 as a new ligand of microfibrils and tropoelastin involved in proper elastic fiber organization.

Keywords: analytical ultracentrifugation; calcium-binding protein; elastin; fibrillin; microfibrillar-associated protein 4; site-directed mutagenesis; surface plasmon resonance (SPR).

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Amino Acid Substitution
Animals
Carrier Proteins / chemistry
Carrier Proteins / genetics
Carrier Proteins / metabolism*
Desmosine / metabolism*
Elastic Tissue / metabolism*
Extracellular Matrix Proteins / chemistry
Extracellular Matrix Proteins / genetics
Extracellular Matrix Proteins / metabolism*
Fibrillin-1
Fibrillins
Glycoproteins / chemistry
Glycoproteins / genetics
Glycoproteins / metabolism*
Humans
Ligands
Male
Mice, Inbred C57BL
Mice, Mutant Strains
Microfibrils / metabolism*
Microfilament Proteins / chemistry
Microfilament Proteins / genetics
Microfilament Proteins / metabolism*
Models, Molecular*
Mutation
Peptide Fragments / chemistry
Peptide Fragments / genetics
Peptide Fragments / metabolism
Protein Interaction Domains and Motifs
Protein Isoforms / chemistry
Protein Isoforms / genetics
Protein Isoforms / metabolism
Protein Multimerization
Protein Transport
Recombinant Proteins / chemistry
Recombinant Proteins / metabolism
Tropoelastin / chemistry
Tropoelastin / genetics
Tropoelastin / metabolism*

Substances

Carrier Proteins
Extracellular Matrix Proteins
FBN1 protein, human
Fbn1 protein, mouse
Fibrillin-1
Fibrillins
Glycoproteins
Ligands
MFAP4 protein, human
MFAP4 protein, mouse
Microfilament Proteins
Peptide Fragments
Protein Isoforms
Recombinant Proteins
Tropoelastin
Desmosine

Associated data

PDB/2J3O