Interaction properties of human mannan-binding lectin (MBL)-associated serine proteases-1 and -2, MBL-associated protein 19, and MBL

N. M. Thielens, S. Cseh, S. Thiel, T. Vorup-Jensen, V. Rossi, J. C. Jensenius, G. J. Arlaud

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Abstract

The mannan-binding lectin (MBL) activation pathway of complement plays an important role in the innate immune defense against pathogenic microorganisms. In human serum, two MBL-associated serine proteases (MASP-1, MASP-2) and MBL-associated protein 19 (MAp19) were found to be associated with MBL. With a view to investigate the interaction properties of these proteins, human MASP-1, MASP-2, MAp19, as well as the N-terminal complement subcomponents Clr/Cls, Uegf, and bone morphogenetic protein-1-epidermal growth factor (CUB-EGF) segments of MASP-1 and MASP-2, were expressed in insect or human kidney cells, and MBL was isolated from human serum. Sedimentation velocity analysis indicated that the MASP-1 and MASP-2 CUB-EGF segments and the homologous protein MAp19 all behaved as homodimers (2.8-3.2 S) in the presence of Ca2+. Although the latter two dimers were not dissociated by EDTA, their physical properties were affected. In contrast, the MASP-1 CUB-EGF homodimer was not sensitive to EDTA. The three proteins and full-length MASP-1 and MASP-2 showed no interaction with each other as judged by gel filtration and surface plasmon resonance spectroscopy. Using the latter technique, MASP-1, MASP-2, their CUB-EGF segments, and MAp19 were each shown to bind to immobilized MBL, with KD values of 0.8 nM (MASP-2), 1.4 nM (MASP-1), 13.0 nM (MAp19 and MASP-2 CUB-EGF), and 25.7 nM (MASP-1 CUB-EGF). The binding was Ca2+-dependent and fully sensitive to EDTA in all cases. These data indicate that MASP-1, MASP-2, and MAp19 each associate as homodimers, and individually form Ca2+-dependent complexes with MBL through the CUB-EGF pair of each protein. This suggests that distinct MBL/MASP complexes may be involved in the activation or regulation of the MBL pathway.

Original languageEnglish
Pages (from-to)5068-5077
Number of pages10
JournalJournal of Immunology
Volume166
Issue number8
Publication statusPublished - Apr 15 2001

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Mannose-Binding Protein-Associated Serine Proteases
Mannose-Binding Lectin
Bone Morphogenetic Protein 1
Proteins
Epidermal Growth Factor
Edetic Acid
human MASP2 protein
Mannose-Binding Lectin Complement Pathway

ASJC Scopus subject areas

  • Immunology

Cite this

Thielens, N. M., Cseh, S., Thiel, S., Vorup-Jensen, T., Rossi, V., Jensenius, J. C., & Arlaud, G. J. (2001). Interaction properties of human mannan-binding lectin (MBL)-associated serine proteases-1 and -2, MBL-associated protein 19, and MBL. Journal of Immunology, 166(8), 5068-5077.

Interaction properties of human mannan-binding lectin (MBL)-associated serine proteases-1 and -2, MBL-associated protein 19, and MBL. / Thielens, N. M.; Cseh, S.; Thiel, S.; Vorup-Jensen, T.; Rossi, V.; Jensenius, J. C.; Arlaud, G. J.

In: Journal of Immunology, Vol. 166, No. 8, 15.04.2001, p. 5068-5077.

Research output: Contribution to journalArticle

Thielens, NM, Cseh, S, Thiel, S, Vorup-Jensen, T, Rossi, V, Jensenius, JC & Arlaud, GJ 2001, 'Interaction properties of human mannan-binding lectin (MBL)-associated serine proteases-1 and -2, MBL-associated protein 19, and MBL', Journal of Immunology, vol. 166, no. 8, pp. 5068-5077.
Thielens, N. M. ; Cseh, S. ; Thiel, S. ; Vorup-Jensen, T. ; Rossi, V. ; Jensenius, J. C. ; Arlaud, G. J. / Interaction properties of human mannan-binding lectin (MBL)-associated serine proteases-1 and -2, MBL-associated protein 19, and MBL. In: Journal of Immunology. 2001 ; Vol. 166, No. 8. pp. 5068-5077.
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abstract = "The mannan-binding lectin (MBL) activation pathway of complement plays an important role in the innate immune defense against pathogenic microorganisms. In human serum, two MBL-associated serine proteases (MASP-1, MASP-2) and MBL-associated protein 19 (MAp19) were found to be associated with MBL. With a view to investigate the interaction properties of these proteins, human MASP-1, MASP-2, MAp19, as well as the N-terminal complement subcomponents Clr/Cls, Uegf, and bone morphogenetic protein-1-epidermal growth factor (CUB-EGF) segments of MASP-1 and MASP-2, were expressed in insect or human kidney cells, and MBL was isolated from human serum. Sedimentation velocity analysis indicated that the MASP-1 and MASP-2 CUB-EGF segments and the homologous protein MAp19 all behaved as homodimers (2.8-3.2 S) in the presence of Ca2+. Although the latter two dimers were not dissociated by EDTA, their physical properties were affected. In contrast, the MASP-1 CUB-EGF homodimer was not sensitive to EDTA. The three proteins and full-length MASP-1 and MASP-2 showed no interaction with each other as judged by gel filtration and surface plasmon resonance spectroscopy. Using the latter technique, MASP-1, MASP-2, their CUB-EGF segments, and MAp19 were each shown to bind to immobilized MBL, with KD values of 0.8 nM (MASP-2), 1.4 nM (MASP-1), 13.0 nM (MAp19 and MASP-2 CUB-EGF), and 25.7 nM (MASP-1 CUB-EGF). The binding was Ca2+-dependent and fully sensitive to EDTA in all cases. These data indicate that MASP-1, MASP-2, and MAp19 each associate as homodimers, and individually form Ca2+-dependent complexes with MBL through the CUB-EGF pair of each protein. This suggests that distinct MBL/MASP complexes may be involved in the activation or regulation of the MBL pathway.",
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AU - Thielens, N. M.

AU - Cseh, S.

AU - Thiel, S.

AU - Vorup-Jensen, T.

AU - Rossi, V.

AU - Jensenius, J. C.

AU - Arlaud, G. J.

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N2 - The mannan-binding lectin (MBL) activation pathway of complement plays an important role in the innate immune defense against pathogenic microorganisms. In human serum, two MBL-associated serine proteases (MASP-1, MASP-2) and MBL-associated protein 19 (MAp19) were found to be associated with MBL. With a view to investigate the interaction properties of these proteins, human MASP-1, MASP-2, MAp19, as well as the N-terminal complement subcomponents Clr/Cls, Uegf, and bone morphogenetic protein-1-epidermal growth factor (CUB-EGF) segments of MASP-1 and MASP-2, were expressed in insect or human kidney cells, and MBL was isolated from human serum. Sedimentation velocity analysis indicated that the MASP-1 and MASP-2 CUB-EGF segments and the homologous protein MAp19 all behaved as homodimers (2.8-3.2 S) in the presence of Ca2+. Although the latter two dimers were not dissociated by EDTA, their physical properties were affected. In contrast, the MASP-1 CUB-EGF homodimer was not sensitive to EDTA. The three proteins and full-length MASP-1 and MASP-2 showed no interaction with each other as judged by gel filtration and surface plasmon resonance spectroscopy. Using the latter technique, MASP-1, MASP-2, their CUB-EGF segments, and MAp19 were each shown to bind to immobilized MBL, with KD values of 0.8 nM (MASP-2), 1.4 nM (MASP-1), 13.0 nM (MAp19 and MASP-2 CUB-EGF), and 25.7 nM (MASP-1 CUB-EGF). The binding was Ca2+-dependent and fully sensitive to EDTA in all cases. These data indicate that MASP-1, MASP-2, and MAp19 each associate as homodimers, and individually form Ca2+-dependent complexes with MBL through the CUB-EGF pair of each protein. This suggests that distinct MBL/MASP complexes may be involved in the activation or regulation of the MBL pathway.

AB - The mannan-binding lectin (MBL) activation pathway of complement plays an important role in the innate immune defense against pathogenic microorganisms. In human serum, two MBL-associated serine proteases (MASP-1, MASP-2) and MBL-associated protein 19 (MAp19) were found to be associated with MBL. With a view to investigate the interaction properties of these proteins, human MASP-1, MASP-2, MAp19, as well as the N-terminal complement subcomponents Clr/Cls, Uegf, and bone morphogenetic protein-1-epidermal growth factor (CUB-EGF) segments of MASP-1 and MASP-2, were expressed in insect or human kidney cells, and MBL was isolated from human serum. Sedimentation velocity analysis indicated that the MASP-1 and MASP-2 CUB-EGF segments and the homologous protein MAp19 all behaved as homodimers (2.8-3.2 S) in the presence of Ca2+. Although the latter two dimers were not dissociated by EDTA, their physical properties were affected. In contrast, the MASP-1 CUB-EGF homodimer was not sensitive to EDTA. The three proteins and full-length MASP-1 and MASP-2 showed no interaction with each other as judged by gel filtration and surface plasmon resonance spectroscopy. Using the latter technique, MASP-1, MASP-2, their CUB-EGF segments, and MAp19 were each shown to bind to immobilized MBL, with KD values of 0.8 nM (MASP-2), 1.4 nM (MASP-1), 13.0 nM (MAp19 and MASP-2 CUB-EGF), and 25.7 nM (MASP-1 CUB-EGF). The binding was Ca2+-dependent and fully sensitive to EDTA in all cases. These data indicate that MASP-1, MASP-2, and MAp19 each associate as homodimers, and individually form Ca2+-dependent complexes with MBL through the CUB-EGF pair of each protein. This suggests that distinct MBL/MASP complexes may be involved in the activation or regulation of the MBL pathway.

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